GPCR BINDING PROTEINS AND SYNTHESIS THEREOF

CROSS-REFERENCE

[0001] This application claims the benefit of U.S. Provisional Patent Application No.

62/556,863 filed on September 11, 2017 which is incorporated herein by reference in its entirety.

BACKGROUND

[0002] G protein-coupled receptors (GPCRs) are implicated in a wide variety of diseases. Raising antibodies to GPCRs has been difficult due to problems in obtaining suitable antigen because GPCRs are often expressed at low levels in cells and are very unstable when purified. Thus, there is a need for improved agents for therapeutic intervention which target GPCRs.

INCORPORATION BY REFERENCE

[0003] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

BRIEF SUMMARY

[0004] Provided herein are antibodies comprising a CDR-H3 comprising a sequence of any one of SEQ ID NOS: 2420 to 2436. Provided herein are antibodies comprising a CDR-H3 comprising a sequence of any one of SEQ ID NOS: 2420 to 2436; and wherein the antibody is a monoclonal antibody, a polyclonal antibody, a bi-specific antibody, a multispecific antibody, a grafted antibody, a human antibody, a humanized antibody, a synthetic antibody, a chimeric antibody, a camelized antibody, a single-chain Fvs (scFv), a single chain antibody, a Fab fragment, a F(ab')2 fragment, a Fd fragment, a Fv fragment, a single-domain antibody, an isolated complementarity determining region (CDR), a diabody, a fragment comprised of only a single monomelic variable domain, disulfide-linked Fvs (sdFv), an intrabody, an anti-idiotypic (anti-Id) antibody, or ab antigen-binding fragments thereof. Provided herein are antibodies wherein the VH domain is IGHV1-18, IGHV1-69, IGHV1-8 IGHV3-21, IGHV3-23, IGHV3- 30/33rn, IGHV3-28, IGHV3-74, IGHV4-39, or IGHV4-59/61. Provided herein are antibodies, wherein the VL domain is IGKVl-39, IGKVl-9, IGKV2-28, IGKV3-11, IGKV3-15, IGKV3-20, IGKV4-1, IGLVl-51, or IGLV2-14. Provided herein are methods of inhibiting GLPIR activity, comprising administering the antibodies as described herein. Provided herein are methods for treatment of a metabolic disorder, comprising administering to a subject in need thereof the antibodies as described herein. In some instances, the antibody comprises a CDR-H3 comprising a sequence of any one of SEQ ID NOS: 2420 to 2436. Provided herein are methods for treatment of a metabolic disorder, wherein the metabolic disorder is Type II diabetes, or obesity. Provided herein are nucleic acids encoding for a protein comprising a sequence of any one of SEQ ID NOS: 2420 to 2436.

[0005] Provided herein are nucleic acid libraries comprising a plurality of nucleic acids, wherein each nucleic acid encodes for a sequence that when translated encodes for an immunoglobulin scaffold, wherein the immunoglobulin scaffold comprises a CDR-H3 loop that comprises a GPCR binding domain, and wherein each nucleic acid comprises a sequence encoding for a sequence variant of the GPCR binding domain. Provided herein are nucleic acid libraries, wherein a length of the CDR-H3 loop is about 20 to about 80 amino acids. Provided herein are nucleic acid libraries, wherein a length of the CDR-H3 loop is about 80 to about 230 base pairs. Provided herein are nucleic acid libraries, wherein the immunoglobulin scaffold further comprises one or more domains selected from variable domain, light chain (VL), variable domain, heavy chain (VH), constant domain, light chain (CL), and constant domain, heavy chain (CH). Provided herein are nucleic acid libraries, wherein the VH domain is IGHVl-18, IGHVl- 69, IGHV1-8 IGHV3-21, IGHV3-23, IGHV3-30/33rn, IGHV3-28, IGHV3-74, IGHV4-39, or IGHV4-59/61. Provided herein are nucleic acid libraries, wherein the VL domain is IGKV1-39, IGKV1-9, IGKV2-28, IGKV3-11, IGKV3-15, IGKV3-20, IGKV4-1, IGLV1-51, or IGLV2-14. Provided herein are nucleic acid libraries, wherein a length of the VH domain is about 90 to about 100 amino acids. Provided herein are nucleic acid libraries, wherein a length of the VL domain is about 90 to about 120 amino acids. Provided herein are nucleic acid libraries, wherein a length of the VH domain is about 280 to about 300 base pairs. Provided herein are nucleic acid libraries, wherein a length of the VL domain is about 300 to about 350 base pairs. Provided herein are nucleic acid libraries, wherein the library comprises at least 105 non-identical nucleic acids. Provided herein are nucleic acid libraries, wherein the immunoglobulin scaffold comprises a single immunoglobulin domain. Provided herein are nucleic acid libraries, wherein the immunoglobulin scaffold comprises a peptide of at most 100 amino acids. Provided herein are vector libraries comprising nucleic acid libraries as described herein. Provided herein are cell libraries comprising nucleic acid libraries as described herein.

[0006] Provided herein are nucleic acid libraries comprising a plurality of nucleic acids, wherein each nucleic acid encodes for a sequence that when translated encodes a GPCR binding domain, and wherein each nucleic acid comprises sequence encoding for a different GPCR binding domain about 20 to about 80 amino acids. Provided herein are nucleic acid libraries, wherein a length of the GPCR binding domain is about 80 to about 230 base pairs. Provided herein are nucleic acid libraries, wherein the GPCR binding domain is designed based on conformational ligand interactions, peptide ligand interactions, small molecule ligand interactions, extracellular domains of GPCRs, or antibodies that target GPCRs. Provided herein are vector libraries comprising nucleic acid libraries as described herein. Provided herein are cell libraries comprising nucleic acid libraries as described herein.

[0007] Provided herein are protein libraries comprising a plurality of proteins, wherein each of the proteins of the plurality of proteins comprise an immunoglobulin scaffold, wherein the immunoglobulin scaffold comprises a CDR-H3 loop that comprises a sequence variant of a GPCR binding domain. Provided herein are protein libraries, wherein a length of the CDR-H3 loop is about 20 to about 80 amino acids. Provided herein are protein libraries, wherein the immunoglobulin scaffold further comprises one or more domains selected from variable domain, light chain (VL), variable domain, heavy chain (VH), constant domain, light chain (CL), and constant domain, heavy chain (CH). Provided herein are protein libraries, wherein the VH domain is IGHV1-18, IGHV1-69, IGHV1-8 IGHV3-21, IGHV3-23, IGHV3-30/33rn, IGHV3- 28, IGHV3-74, IGHV4-39, or IGHV4-59/61. Provided herein are protein libraries, wherein the VL domain is IGKVl-39, IGKVl-9, IGKV2-28, IGKV3-11, IGKV3-15, IGKV3-20, IGKV4-1, IGLV1-51, or IGLV2-14. Provided herein are protein libraries, wherein a length of the VH domain is about 90 to about 100 amino acids. Provided herein are protein libraries, wherein a length of the VL domain is about 90 to about 120 amino acids. Provided herein are protein libraries, wherein the plurality of proteins is used to generate a peptidomimetic library. Provided herein are protein libraries, wherein the protein library comprises peptides. Provided herein are protein libraries, wherein the protein library comprises immunoglobulins. Provided herein are protein libraries, wherein the protein library comprises antibodies. Provided herein are cell libraries comprising protein libraries as described herein.

[0008] Provided herein are protein libraries comprising a plurality of proteins, wherein the plurality of proteins comprises sequence encoding for different GPCR binding domains, and wherein the length of each GPCR binding domain is about 20 to about 80 amino acids. Provided herein are protein libraries, wherein the protein library comprises peptides. Provided herein are protein libraries, wherein the protein library comprises immunoglobulins. Provided herein are protein libraries, wherein the protein library comprises antibodies. Provided herein are protein libraries, wherein the plurality of proteins are used to generate a peptidomimetic library.

Provided herein are cell libraries comprising protein libraries as described herein.

[0009] Provided herein are vector libraries comprising a nucleic acid library described herein. Provided herein are cell libraries comprising a nucleic acid library described herein. Provided herein are cell libraries comprising a protein library described herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 depicts a schematic of G protein-coupled receptor (GPCR) ligand interaction surfaces.

[0011] Figure 2A depicts a first schematic of an immunoglobulin scaffold.

[0012] Figure 2B depicts a second schematic of an immunoglobulin scaffold.

[0013] Figure 3 depicts a schematic of a motif for placement in a scaffold.

[0014] Figure 4 depicts a schematic of a GPCR.

[0015] Figure 5 depicts schematics of segments for assembly of clonal fragments and non- clonal fragments.

[0016] Figure 6 depicts schematics of segments for assembly of clonal fragments and non- clonal fragments.

[0017] Figure 7 presents a diagram of steps demonstrating an exemplary process workflow for gene synthesis as disclosed herein.

[0018] Figure 8 illustrates an example of a computer system.

[0019] Figure 9 is a block diagram illustrating an architecture of a computer system.

[0020] Figure 10 is a diagram demonstrating a network configured to incorporate a plurality of computer systems, a plurality of cell phones and personal data assistants, and Network Attached Storage (NAS).

[0021] Figure 11 is a block diagram of a multiprocessor computer system using a shared virtual address memory space.

[0022] Figures 12A-12C depict sequences of immunoglobulin scaffolds.

[0023] Figure 13 depicts sequences of G protein-coupled receptors scaffolds.

[0024] Figure 14 is a graph of normalized reads for a library for variable domain, heavy chains.

[0025] Figure 15 is a graph of normalized reads for a library for variable domain, light chains.

[0026] Figure 16 is a graph of normalized reads for a library for heavy chain

complementarity determining region 3.

[0027] Figure 17A is a plot of light chain frameworks assayed for folding.

[0028] Figure 17B is a plot of light chain frameworks assayed for thermostability.

[0029] Figure 17C is a plot of light chain frameworks assayed for motif display using FLAG tag.

[0030] Figure 17D is a plot of light chain frameworks assayed for motif display using His tag. [0031] Figure 18A is a plot of heavy chain frameworks assayed for folding.

[0032] Figure 18B is a plot of heavy chain frameworks assayed for stability.

[0033] Figure 18C is a plot of heavy chain frameworks assayed for motif display using

FLAG tag.

[0034] Figure 18D is a plot of heavy chain frameworks assayed for motif display using His tag.

[0035] Figure 18E is a plot of heavy chain frameworks assayed for expression.

[0036] Figure 18F is a plot of heavy chain frameworks assayed for selection specificity.

[0037] Figures 19A-19C depict images of G protein-coupled receptors visualized by fluorescent antibodies.

[0038] Figures 20A-20C depict images of G protein-coupled receptors visualized by auto- fluorescent proteins.

[0039] Figure 21A depicts a schematic of an immunoglobulin scaffold comprising a VH domain attached to a VL domain using a linker.

[0040] Figure 21B depicts a schematic of a full-domain architecture of an immunoglobulin scaffold comprising a VH domain attached to a VL domain using a linker, a leader sequence, and pill sequence.

[0041] Figure 21C depicts a schematic of four framework elements (FW1, FW2, FW3, FW4) and the variable 3 CDR (LI, L2, L3) elements for a VL or VH domain.

DETAILED DESCRIPTION

[0042] The present disclosure employs, unless otherwise indicated, conventional molecular biology techniques, which are within the skill of the art. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art.

[0043] Definitions

[0044] Throughout this disclosure, various embodiments are presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of any embodiments.

Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range to the tenth of the unit of the lower limit unless the context clearly dictates otherwise. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual values within that range, for example, 1.1, 2, 2.3, 5, and 5.9. This applies regardless of the breadth of the range. The upper and lower limits of these intervening ranges may independently be included in the smaller ranges, and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure, unless the context clearly dictates otherwise.

[0045] The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of any embodiment. As used herein, the singular forms "a," "an" and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise. It will be further understood that the terms "comprises" and/or

"comprising," when used in this specification, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items.

[0046] Unless specifically stated or obvious from context, as used herein, the term "about" in reference to a number or range of numbers is understood to mean the stated number and numbers +/- 10% thereof, or 10% below the lower listed limit and 10% above the higher listed limit for the values listed for a range.

[0047] Unless specifically stated, as used herein, the term "nucleic acid" encompasses double- or triple-stranded nucleic acids, as well as single-stranded molecules. In double- or triple -stranded nucleic acids, the nucleic acid strands need not be coextensive (i.e., a double- stranded nucleic acid need not be double-stranded along the entire length of both strands).

Nucleic acid sequences, when provided, are listed in the 5 ' to 3 ' direction, unless stated otherwise. Methods described herein provide for the generation of isolated nucleic acids.

Methods described herein additionally provide for the generation of isolated and purified nucleic acids. A "nucleic acid" as referred to herein can comprise at least 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 600, 700, 800, 900, 1000, 1 100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, or more bases in length. Moreover, provided herein are methods for the synthesis of any number of polypeptide-segments encoding nucleotide sequences, including sequences encoding non- ribosomal peptides (NRPs), sequences encoding non-ribosomal peptide-synthetase (NRPS) modules and synthetic variants, polypeptide segments of other modular proteins, such as antibodies, polypeptide segments from other protein families, including non-coding DNA or RNA, such as regulatory sequences e.g. promoters, transcription factors, enhancers, siRNA, shRNA, R Ai, miRNA, small nucleolar RNA derived from microRNA, or any functional or structural DNA or RNA unit of interest. The following are non-limiting examples of polynucleotides: coding or non-coding regions of a gene or gene fragment, intergenic DNA, loci (locus) defined from linkage analysis, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, short interfering RNA (siRNA), short-hairpin RNA (shRNA), micro-RNA (miRNA), small nucleolar RNA, ribozymes, complementary DNA (cDNA), which is a DNA representation of mRNA, usually obtained by reverse transcription of messenger RNA (mRNA) or by amplification; DNA molecules produced synthetically or by amplification, genomic DNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers. cDNA encoding for a gene or gene fragment referred herein may comprise at least one region encoding for exon sequences without an intervening intron sequence in the genomic equivalent sequence.

[0048] GPCR Libraries

[0049] Provided herein are methods and compositions relating to G protein-coupled receptor (GPCR) binding libraries comprising nucleic acids encoding for a scaffold comprising a GPCR binding domain. Scaffolds as described herein can stably support a GPCR binding domain. The GPCR binding domain may be designed based on surface interactions of a GPCR ligand and the GPCR. Libraries as described herein may be further variegated to provide for variant libraries comprising nucleic acids each encoding for a predetermined variant of at least one predetermined reference nucleic acid sequence. Further described herein are protein libraries that may be generated when the nucleic acid libraries are translated. In some instances, nucleic acid libraries as described herein are transferred into cells to generate a cell library. Also provided herein are downstream applications for the libraries synthesized using methods described herein.

Downstream applications include identification of variant nucleic acids or protein sequences with enhanced biologically relevant functions, e.g., improved stability, affinity, binding, functional activity, and for the treatment or prevention of a disease state associated with GPCR signaling.

[0050] Scaffold Libraries

[0051] Provided herein are libraries comprising nucleic acids encoding for a scaffold, wherein sequences for GPCR binding domains are placed in the scaffold. Scaffold described herein allow for improved stability for a range of GPCR binding domain encoding sequences when inserted into the scaffold, as compared to an unmodified scaffold. Exemplary scaffolds include, but are not limited to, a protein, a peptide, an immunoglobulin, derivatives thereof, or combinations thereof. In some instances, the scaffold is an immunoglobulin. Scaffolds as described herein comprise improved functional activity, structural stability, expression, specificity, or a combination thereof. In some instances, scaffolds comprise long regions for supporting a GPCR binding domain.

[0052] Provided herein are libraries comprising nucleic acids encoding for a scaffold, wherein the scaffold is an immunoglobulin. In some instances, the immunoglobulin is an antibody. As used herein, the term antibody will be understood to include proteins having the characteristic two-armed, Y-shape of a typical antibody molecule as well as one or more fragments of an antibody that retain the ability to specifically bind to an antigen. Exemplary antibodies include, but are not limited to, a monoclonal antibody, a polyclonal antibody, a bi- specific antibody, a multispecific antibody, a grafted antibody, a human antibody, a humanized antibody, a synthetic antibody, a chimeric antibody, a camelized antibody, a single-chain Fvs (scFv) (including fragments in which the VL and VH are joined using recombinant methods by a synthetic or natural linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules, including single chain Fab and scFab), a single chain antibody, a Fab fragment (including monovalent fragments comprising the VL, VH, CL, and CHI domains), a F(ab')2 fragment (including bivalent fragments comprising two Fab fragments linked by a disulfide bridge at the hinge region), a Fd fragment (including fragments comprising the VH and CHI fragment), a Fv fragment (including fragments comprising the VL and VH domains of a single arm of an antibody), a single-domain antibody (dAb or sdAb) (including fragments comprising a VH domain), an isolated complementarity determining region (CDR), a diabody (including fragments comprising bivalent dimers such as two VL and VH domains bound to each other and recognizing two different antigens), a fragment comprised of only a single monomeric variable domain, disulfide-linked Fvs (sdFv), an intrabody, an anti- idiotypic (anti-Id) antibody, or ab antigen-binding fragments thereof. In some instances, the libraries disclosed herein comprise nucleic acids encoding for a scaffold, wherein the scaffold is a Fv antibody, including Fv antibodies comprised of the minimum antibody fragment which contains a complete antigen-recognition and antigen-binding site. In some embodiments, the Fv antibody consists of a dimer of one heavy chain and one light chain variable domain in tight, non-covalent association, and the three hypervariable regions of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. In some embodiments, the six hypervariable regions confer antigen-binding specificity to the antibody. In some embodiments, a single variable domain (or half of an Fv comprising only three hypervariable regions specific for an antigen, including single domain antibodies isolated from camelid animals comprising one heavy chain variable domain such as VHH antibodies or nanobodies) has the ability to recognize and bind antigen. In some instances, the libraries disclosed herein comprise nucleic acids encoding for a scaffold, wherein the scaffold is a single-chain Fv or scFv, including antibody fragments comprising a VH, a VL, or both a VH and VL domain, wherein both domains are present in a single polypeptide chain. In some embodiments, the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains allowing the scFv to form the desired structure for antigen binding. In some instances, a scFv is linked to the Fc fragment or a VHH is linked to the Fc fragment (including minibodies). In some instances, the antibody comprises immunoglobulin molecules and immunologically active fragments of immunoglobulin molecules, e.g., molecules that contain an antigen binding site. Immunoglobulin molecules are of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgG 1, IgG 2, IgG 3, IgG 4, IgA 1 and IgA 2) or subclass.

[0053] Libraries described herein comprising nucleic acids encoding for a scaffold, wherein the scaffold is an immunoglobulin, comprise variations in at least one region of the

immunoglobulin. Exemplary regions of the antibody for variation include, but are not limited to, a complementarity-determining region (CDR), a variable domain, or a constant domain. In some instances, the CDR is CDR1, CDR2, or CDR3. In some instances, the CDR is a heavy domain including, but not limited to, CDR-H1, CDR-H2, and CDR-H3. In some instances, the CDR is a light domain including, but not limited to, CDR-L1, CDR-L2, and CDR-L3. In some instances, the variable domain is variable domain, light chain (VL) or variable domain, heavy chain (VH). In some instances, the VL domain comprises kappa or lambda chains. In some instances, the constant domain is constant domain, light chain (CL) or constant domain, heavy chain (CH).

[0054] Methods described herein provide for synthesis of libraries comprising nucleic acids encoding for a scaffold, wherein each nucleic acid encodes for a predetermined variant of at least one predetermined reference nucleic acid sequence. In some cases, the predetermined reference sequence is a nucleic acid sequence encoding for a protein, and the variant library comprises sequences encoding for variation of at least a single codon such that a plurality of different variants of a single residue in the subsequent protein encoded by the synthesized nucleic acid are generated by standard translation processes. In some instances, the scaffold library comprises varied nucleic acids collectively encoding variations at multiple positions. In some instances, the variant library comprises sequences encoding for variation of at least a single codon of a CDR- Hl, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR-L3, VL, or VH domain. In some instances, the variant library comprises sequences encoding for variation of multiple codons of a CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR-L3, VL, or VH domain. In some instances, the variant library comprises sequences encoding for variation of multiple codons of framework element 1 (FWl), framework element 2 (FW2), framework element 3 (FW3), or framework element 4 (FW4). An exemplary number of codons for variation include, but are not limited to, at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 225, 250, 275, 300, or more than 300 codons.

[0055] In some instances, the at least one region of the immunoglobulin for variation is from heavy chain V-gene family, heavy chain D-gene family, heavy chain J-gene family, light chain V-gene family, or light chain J-gene family. In some instances, the light chain V-gene family comprises immunoglobulin kappa (IGK) gene or immunoglobulin lambda (IGL). Exemplary genes include, but are not limited to, IGHV1-18, IGHV1-69, IGHV1-8, IGHV3-21, IGHV3-23, IGHV3-30/33rn, IGHV3-28, IGHV1-69, IGHV3-74, IGHV4-39, IGHV4-59/61, IGKV1-39, IGKV1-9, IGKV2-28, IGKV3-11, IGKV3-15, IGKV3-20, IGKV4-1, IGLV1-51, and IGLV2-14.

[0056] Provided herein are libraries comprising nucleic acids encoding for immunoglobulin scaffolds, wherein the libraries are synthesized with various numbers of fragments. In some instances, the fragments comprise the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR- L3, VL, or VH domain. In some instances, the fragments comprise framework element 1 (FWl), framework element 2 (FW2), framework element 3 (FW3), or framework element 4 (FW4). In some instances, the scaffold libraries are synthesized with at least or about 2 fragments, 3 fragments, 4 fragments, 5 fragments, or more than 5 fragments. The length of each of the nucleic acid fragments or average length of the nucleic acids synthesized may be at least or about 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, 600, or more than 600 base pairs. In some instances, the length is about 50 to 600, 75 to 575, 100 to 550, 125 to 525, 150 to 500, 175 to 475, 200 to 450, 225 to 425, 250 to 400, 275 to 375, or 300 to 350 base pairs.

[0057] Libraries comprising nucleic acids encoding for immunoglobulin scaffolds as described herein comprise various lengths of amino acids when translated. In some instances, the length of each of the amino acid fragments or average length of the amino acid synthesized may be at least or about 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, or more than 150 amino acids. In some instances, the length of the amino acid is about 15 to 150, 20 to 145, 25 to 140, 30 to 135, 35 to 130, 40 to 125, 45 to 120, 50 to 115, 55 to 110, 60 to 110, 65 to 105, 70 to 100, or 75 to 95 amino acids. In some instances, the length of the amino acid is about 22 amino acids to about 75 amino acids. In some instances, the immunoglobulin scaffolds comprise at least or about 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, or more than 5000 amino acids. [0058] A number of variant sequences for the at least one region of the immunoglobulin for variation are de novo synthesized using methods as described herein. In some instances, a number of variant sequences is de novo synthesized for CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR-L3, VL, VH, or combinations thereof. In some instances, a number of variant sequences is de novo synthesized for framework element 1 (FW1), framework element 2 (FW2), framework element 3 (FW3), or framework element 4 (FW4). The number of variant sequences may be at least or about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95,

100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, or more than 500 sequences. In some instances, the number of variant sequences is at least or about 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, or more than 8000 sequences. In some instances, the number of variant sequences is about 10 to 500, 25 to 475, 50 to 450, 75 to 425, 100 to 400, 125 to 375, 150 to 350, 175 to 325, 200 to 300, 225 to 375, 250 to 350, or 275 to 325 sequences.

[0059] Variant sequences for the at least one region of the immunoglobulin, in some instances, vary in length or sequence. In some instances, the at least one region that is de novo synthesized is for CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR-L3, VL, VH, or combinations thereof. In some instances, the at least one region that is de novo synthesized is for framework element 1 (FW1), framework element 2 (FW2), framework element 3 (FW3), or framework element 4 (FW4). In some instances, the variant sequence comprises at least or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, or more than 50 variant nucleotides or amino acids as compared to wild-type. In some instances, the variant sequence comprises at least or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, or 50 additional nucleotides or amino acids as compared to wild-type. In some instances, the variant sequence comprises at least or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, or 50 less nucleotides or amino acids as compared to wild-type. In some instances, the libraries comprise at least or about

10 ¹ , 10 ² , 10 ³ , 10 ⁴ , 10 ⁵ , 10 ⁶ , 10 ⁷ , 10 ⁸ , 10 ⁹ , 10 ¹⁰ , or more than 10 ¹⁰ variants.

[0060] Following synthesis of scaffold libraries, scaffold libraries may be used for screening and analysis. For example, scaffold libraries are assayed for library displayability and panning. In some instances, displayability is assayed using a selectable tag. Exemplary tags include, but are not limited to, a radioactive label, a fluorescent label, an enzyme, a chemiluminescent tag, a colorimetric tag, an affinity tag or other labels or tags that are known in the art. In some instances, the tag is histidine, polyhistidine, myc, hemagglutinin (HA), or FLAG. In some instances, scaffold libraries are assayed by sequencing using various methods including, but not limited to, single-molecule real-time (SMRT) sequencing, Polony sequencing, sequencing by ligation, reversible terminator sequencing, proton detection sequencing, ion semiconductor sequencing, nanopore sequencing, electronic sequencing, pyrosequencing, Maxam-Gilbert sequencing, chain termination (e.g., Sanger) sequencing, +S sequencing, or sequencing by synthesis.

[0061] In some instances, the scaffold libraries are assayed for functional activity, structural stability (e.g., thermal stable or pH stable), expression, specificity, or a combination thereof. In some instances, the scaffold libraries are assayed for scaffolds capable of folding. In some instances, a region of the antibody is assayed for functional activity, structural stability, expression, specificity, folding, or a combination thereof. For example, a VH region or VL region is assayed for functional activity, structural stability, expression, specificity, folding, or a combination thereof.

[0062] GPCR Libraries

[0063] Provided herein are G protein-coupled receptor (GPCR) binding libraries comprising nucleic acids encoding for scaffolds comprising sequences for GPCR binding domains. In some instances, the scaffolds are immunoglobulins. In some instances, the scaffolds comprising sequences for GPCR binding domains are determined by interactions between the GPCR binding domains and the GPCRs.

[0064] Provided herein are libraries comprising nucleic acids encoding scaffolds comprising GPCR binding domains, wherein the GPCR binding domains are designed based on surface interactions on the GPCRs. Exemplary GPCRs are seen in Table 1. In some instances, the GPCR binding domains interact with the amino- or carboxy-terminus of the GPCR. In some instances, the GPCR binding domains interact with at least one transmembrane domain including, but not limited to, transmembrane domain 1 (TMl), transmembrane domain 2 (TM2), transmembrane domain 3 (TM3), transmembrane domain 4 (TM4), transmembrane domain 5 (TM5), transmembrane domain 6 (TM6), and transmembrane domain 7 (TM7). In some instances, the GPCR binding domains interact with an intracellular surface of the GPCR. For example, the GPCR binding domains interact with at least one intracellular loop including, but not limited to, intracellular loop 1 (ICL1), intracellular loop 2 (ICL2), and intracellular loop 3 (ICL3). In some instances, the GPCR binding domains interact with an extracellular surface of the GPCR. See FIG. 1. For example, the GPCR binding domains interact with at least one extracellular domain (ECD) or extracellular loop (ECL) of the GPCR. The extracellular loops include, but are not limited to, extracellular loop 1 (ECL1), extracellular loop 2 (ECL2), and extracellular loop 3 (ECL3). Table 1. List of GPCRs

GPCR Gene Name Accession

Number

5-hydroxytryptamine receptor 1A HTR1A P08908

5-hydroxytryptamine receptor IB HTR1B P28222

5-hydroxytryptamine receptor ID HTR1D P28221

5-hydroxytryptamine receptor IE HTR1E P28566

5-hydroxytryptamine receptor IF HTR1F P30939

5-hydroxytryptamine receptor 2 A HTR2A P28223

5-hydroxytryptamine receptor 2B HTR2B P41595

5-hydroxytryptamine receptor 2C HTR2C P28335

5-hydroxytryptamine receptor 4 HTR4 Q13639

5-hydroxytryptamine receptor 5 A HTR5A P47898

5-hydroxytryptamine receptor 6 HTR6 P50406

5-hydroxytryptamine receptor HTR7 P34969

Adenosine receptor Al ADORA1 P30542

Adenosine receptor A2a ADORA2A P29274

Adenosine receptor A2b ADORA2B P29275

Adenosine A3 receptor ADORA3 P33765

Muscarinic acetylcholine receptor Ml CHRM1 PI 1229

Muscarinic acetylcholine receptor M2 CHRM2 P08172

Muscarinic acetylcholine receptor M3 CHRM3 P20309

Muscarinic acetylcholine receptor M4 CHRM4 P08173

Muscarinic acetylcholine receptor M5 CHRM5 P08912

Adrenocorticotropic hormone receptor MC2R Q01718 a-lA adrenergic receptor ADRA1A P35348 a- IB adrenergic receptor ADRA1B P35368 a- ID adrenergic receptor ADRA1D P25100 a-2A adrenergic receptor ADRA2A P08913 a-2B adrenergic receptor ADRA2B P18089 a-2C adrenergic receptor ADRA2C P18825 β-l adrenergic receptor ADRB1 P08588 β-2 adrenergic receptor ADRB2 P07550 β-3 adrenergic receptor ADRB3 P13945

Type-1 angiotensin II receptor AGTR1 P30556

Duffy antigen/chemokine receptor DARC Q16570

Endothelin-1 receptor EDNRA P25101

Endothelin B receptor EDNRB P24530

N-formyl peptide receptor 2 FPR2 P25090

Follicle-stimulating hormone receptor FSHR P23945

Galanin receptor type 1 GALR1 P47211

Galanin receptor type 2 GALR2 043603

Galanin receptor type 3 GALR3 060755

Gastrin/cholecystokinin type B receptor CCKBR P32239

Gonadotropin-releasing hormone receptor GNRHR P30968

Putative gonadotropin-releasing hormone II GNRHR2 Q96P88 receptor G-protein coupled oestrogen receptor 1 GPER Q99527

Uracil nucleotide/cysteinyl leukotriene receptor GPR17 013304

Putative G-protein coupled receptor 44 GPR44 Q9Y5Y4

G-protein coupled receptor 55 GPR55 Q9Y2T6

Gastrin-releasing peptide receptor GRPR P30550

Histamine HI receptor HRHl P35367

Histamine H2 receptor HRH2 P25021

Histamine H3 receptor HRH3 Q9Y5N1

Histamine H4 receptor HRH4 Q9H3N8

KiSS-1 receptor KISS1R Q969F8

Lysophosphatidic acid receptor 1 LPAR1 Q92633

Lysophosphatidic acid receptor 2 LPAR2 Q9HBW0

Lysophosphatidic acid receptor 3 LPAR3 Q9UBY5

Lysophosphatidic acid receptor 4 LPAR4 099677

Lysophosphatidic acid receptor 6 LPAR6 P43657

Lutropin-choriogonadotropic hormone receptor LHCGR P22888

Leukotriene B4 receptor 1 LTB4R Q15722

Leukotriene B4 receptor 2 LTB4R2 Q9NPC1

Melanocortin receptor 3 MC3R P41968

Melanocortin receptor 4 MC4R P32245

Melanocortin receptor 5 MC5R P33032

Olfactory receptor 10G9 OR10G9 Q8NGN4

Olfactory receptor 10H1 OR10H1 Q9Y4A9

Olfactory receptor 10H2 OR10H2 060403

Olfactory receptor 10H3 OR10H3 060404

Olfactory receptor 10H4 OR10H4 Q8NGA5

Olfactory receptor 10H5 OR10H5 Q8NGA6

Olfactory receptor 10J1 OR10J1 P30954

Olfactory receptor 10J3 OR10J3 Q5JRS4

Olfactory receptor 10J5 OR10J5 Q8NHC4

Olfactory receptor 10K1 OR10K1 Q8NGX5

Olfactory receptor 10K2 OR10K2 Q6IF99

Olfactory receptor 10P1 OR10P1 Q8NGE3

Olfactory receptor 10Q 1 OR10Q1 Q8NGQ4

Olfactory receptor 10R2 OR10R2 Q8NGX6

Olfactory receptor 10S 1 OR10S 1 Q8NGN2

Olfactory receptor 10T2 OR10T2 Q8NGX3

Olfactory receptor 10V1 OR10V1 Q8NGI7

Olfactory receptor 10W1 OR10W1 Q8NGF6

Olfactory receptor 14A2 OR14A2 Q96R54

Olfactory receptor 14C36 OR14C36 Q8NHC7

Olfactory receptor 1411 OR14I1 A6ND48

Olfactory receptor 14J1 OR14J1 Q9UGF5

Olfactory receptor 14K1 OR14K1 Q8NGZ2

Olfactory receptor 2A12 OR2A12 Q8NGT7

Olfactory receptor 2A14 OR2A14 Q96R47

Olfactory receptor 2A25 OR2A25 A4D2G3

Olfactory receptor 2AG1 OR2AG1 Q9H205 Olfactory receptor 2AG2 OR2AG2 A6NM03

Olfactory receptor 2AJ1 OR2AJ1 Q8NGZ0

Olfactory receptor 2AK2 OR2AK2 Q8NG84

Olfactory receptor 2AP1 OR2AP1 Q8NGE2

Olfactory receptor 2AT4 OR2AT4 A6N D4

Olfactory receptor 5112 OR51I2 Q9H344

Olfactory receptor 51 Jl OR51J1 Q9H342

Olfactory receptor 51L1 OR51L1 Q8NGJ5

Olfactory receptor 51M1 OR51M1 Q9H341

Olfactory receptor 51Q 1 OR51Q1 Q8NH59

Olfactory receptor 51 S 1 OR51S 1 Q8NGJ8

Olfactory receptor 51T1 OR51T1 Q8NGJ9

Olfactory receptor 51V1 OR51V1 Q9H2C8

Olfactory receptor 52A 1 OR52A1 Q9UKL2

Olfactory receptor 52A5 OR52A5 Q9H2C5

Olfactory receptor 52B2 OR52B2 Q96RD2

Olfactory receptor 52B4 OR52B4 Q8NGK2

Olfactory receptor 52B6 OR52B6 Q8NGF0

Olfactory receptor 52D 1 OR52D1 Q9H346

Olfactory receptor 52E2 OR52E2 Q8NGJ4

Olfactory receptor 52E4 OR52E4 Q8NGH9

Olfactory receptor 52E5 OR52E5 Q8NH55

Olfactory receptor 52E6 OR52E6 Q96RD3

Olfactory receptor 52E8 OR52E8 Q6IFG1

Olfactory receptor 52H1 OR52H1 Q8NGJ2

Olfactory receptor 5211 OR52I1 Q8NGK6

Olfactory receptor 5212 OR52I2 Q8NH67

Olfactory receptor 52K1 OR52K1 Q8NGK4

Olfactory receptor 52K2 OR52K2 Q8NGK3

Olfactory receptor 52L1 OR52L1 Q8NGH7

Olfactory receptor 52M1 OR52M1 Q8NGK5

Olfactory receptor 52N 1 OR52N1 Q8NH53

Olfactory receptor 52N2 OR52N2 Q8NGI0

Olfactory receptor 52N4 OR52N4 Q8NGI2

Olfactory receptor 52N5 OR52N5 Q8NH56

Olfactory receptor 52R1 OR52R1 Q8NGF1

Olfactory receptor 52W1 OR52W1 Q6IF63

Red-sensitive opsin OPN1LW P04000

Visual pigment-like receptor peropsin RRH 014718

Olfactory receptor 1 A 1 OR1A1 Q9P1Q5

Olfactory receptor 1A2 OR1A2 Q9Y585

Olfactory receptor 1B 1 OR1B1 Q8NGR6

Olfactory receptor 1C1 OR1C1 Q15619

Olfactory receptor 1D2 OR1D2 P34982

Olfactory receptor 1F1 OR1F1 043749

Olfactory receptor IF 12 OR1F12 Q8NHA8

Olfactory receptor 1G1 OR1G1 P47890

Olfactory receptor 111 OR1I1 060431 Olfactory receptor 1J1 OR1J1 Q8NGS3

Olfactory receptor 1J2 OR1J2 Q8NGS2

Olfactory receptor 1J4 OR1J4 Q8NGS 1

Olfactory receptor 1K1 OR1K1 Q8NGR3

Olfactory receptor 1L1 OR1L1 Q8NH94

Olfactory receptor 1L3 OR1L3 Q8NH93

Olfactory receptor 1L4 OR1L4 Q8NGR5

Olfactory receptor 1L6 OR1L6 Q8NGR2

Olfactory receptor 1L8 OR1L8 Q8NGR8

Olfactory receptor 1M1 OR1M1 Q8NGA1

Olfactory receptor IN 1 OR1N1 Q8NGS0

Olfactory receptor 1N2 OR1N2 Q8NGR9

Olfactory receptor 1Q 1 OR1Q1 Q15612

Olfactory receptor 1S 1 OR1S1 Q8NH92

Olfactory receptor 1S2 OR1S2 Q8NGQ3

Olfactory receptor 2A2 OR2A2 Q6IF42

Olfactory receptor 2A4 OR2A4 095047

Olfactory receptor 2B2 OR2B2 Q9GZK3

Putative olfactory receptor 2B3 OR2B3 076000

Olfactory receptor 2B6 OR2B6 P58173

Putative olfactory receptor 2B8 OR2B8P P59922

Olfactory receptor 2T5 OR2T5 Q6IEZ7

Olfactory receptor 2T6 OR2T6 Q8NHC8

Olfactory receptor 2T8 OR2T8 A6NH00

Olfactory receptor 2V 1 OR2V1 Q8NHB 1

Olfactory receptor 2V2 OR2V2 Q96R30

Olfactory receptor 2W1 OR2W1 Q9Y3N9

Olfactory receptor 2W3 OR2W3 Q7Z3T1

Olfactory receptor 2Y 1 OR2Y1 Q8NGV0

Olfactory receptor 2Z 1 OR2Z1 Q8NG97

Olfactory receptor 3 A 1 OR3A1 P47881

Olfactory receptor 3A2 OR3A2 P47893

Olfactory receptor 3A3 OR3A3 P47888

Olfactory receptor 3A4 OR3A4 P47883

Putative olfactory receptor 4A4 OR4A4P Q8NGN8

Olfactory receptor 4A5 OR4A5 Q8NH83

Olfactory receptor 4A8 OR4A8P P0C604

Olfactory receptor 4B 1 OR4B1 Q8NGF8

Olfactory receptor 4C3 OR4C3 Q8NH37

Olfactory receptor 4C5 OR4C5 Q8NGB2

Olfactory receptor 4C6 OR4C6 Q8NH72

Olfactory receptor 4C 11 OR4C11 Q6IEV9

Olfactory receptor 4C12 OR4C12 Q96R67

Olfactory receptor 4C13 OR4C13 Q8NGP0

Olfactory receptor 4C15 OR4C15 Q8NGM1

Olfactory receptor 4C16 OR4C16 Q8NGL9

Olfactory receptor 4D 1 OR4D1 Q15615

Olfactory receptor 4D2 OR4D2 P58180 Olfactory receptor 4D5 OR4D5 Q8NGN0

Olfactory receptor 4D6 OR4D6 Q8NGJ1

Olfactory receptor 4D9 OR4D9 Q8NGE8

Olfactory receptor 4D 10 OR4D10 Q8NGI6

Olfactory receptor 4D 11 OR4D11 Q8NGI4

Olfactory receptor 5B 17 OR5B17 Q8NGF7

Olfactory receptor 5B21 OR5B21 A6NL26

Olfactory receptor 5C1 OR5C1 Q8NGR4

Olfactory receptor 5D 13 OR5D13 Q8NGL4

Olfactory receptor 5D 14 OR5D14 Q8NGL3

Olfactory receptor 5D 16 OR5D16 Q8NGK9

Olfactory receptor 5D 18 OR5D18 Q8NGL1

Olfactory receptor 5F 1 OR5F1 095221

Olfactory receptor 5H1 OR5H1 A6NKK0

Olfactory receptor 5H2 OR5H2 Q8NGV7

Olfactory receptor 5H6 OR5H6 Q8NGV6

Olfactory receptor 511 OR5I1 Q13606

Olfactory receptor 5J2 OR5J2 Q8NH18

Olfactory receptor 5K1 OR5K1 Q8NHB7

Olfactory receptor 5K2 OR5K2 Q8NHB8

Olfactory receptor 5K3 OR5K3 A6NET4

Olfactory receptor 5K4 OR5K4 A6NMS3

Olfactory receptor 5L1 OR5L1 Q8NGL2

Olfactory receptor 5L2 OR5L2 Q8NGL0

Olfactory receptor 5M1 OR5M1 Q8NGP8

Olfactory receptor 5M3 OR5M3 Q8NGP4

Olfactory receptor 5M8 OR5M8 Q8NGP6

Olfactory receptor 5M9 OR5M9 Q8NGP3

Olfactory receptor 5M10 OR5M10 Q6IEU7

Olfactory receptor 5M11 OR5M11 Q96RB7

Olfactory receptor 5P2 OR5P2 Q8WZ92

Olfactory receptor 5P3 OR5P3 Q8WZ94

Olfactory receptor 5R1 OR5R1 Q8NH85

Olfactory receptor 5T1 OR5T1 Q8NG75

Olfactory receptor 5T2 OR5T2 Q8NGG2

Olfactory receptor 5T3 OR5T3 Q8NGG3

Olfactory receptor 5W2 OR5W2 Q8NH69

Olfactory receptor 7G3 OR7G3 Q8NG95

Olfactory receptor 8A 1 OR8A1 Q8NGG7

Olfactory receptor 8B3 OR8B3 Q8NGG8

Olfactory receptor 8B4 OR8B4 Q96RC9

Olfactory receptor 8B8 OR8B8 Q15620

Olfactory receptor 8B 12 OR8B12 Q8NGG6

Olfactory receptor 8D 1 OR8D1 Q8WZ84

Olfactory receptor 8D2 OR8D2 Q9GZM6

Olfactory receptor 8D4 OR8D4 Q8NGM9

Orexin receptor type 1 HCRTR1 043613

Orexin receptor type 2 HCRTR2 043614 Oxoeicosanoid receptor 1 OXER1 Q8TDS5

Oxytocin receptor OXTR P30559

P2Y purinoceptor 1 P2RY1 P47900

P2Y purinoceptor 2 P2RY2 P41231

P2Y purinoceptor 4 P2RY4 P51582

P2Y purinoceptor 6 P2RY6 015077

P2Y purinoceptor 8 P2RY8 Q86VZ1

Putative P2Y purinoceptor 10 P2RY10 000398

P2Y purinoceptor 11 P2RY11 Q96G91

P2Y purinoceptor 12 P2RY12 Q9H244

P2Y purinoceptor 13 P2RY13 Q9BPV8

P2Y purinoceptor 14 P2RY14 Q15391

Proteinase-activated receptor 1 F2R P251 16

Proteinase-activated receptor 2 F2RL1 P55085

Proteinase-activated receptor 3 F2RL2 000254

Proteinase-activated receptor 4 F2RL3 Q96RI0

Prostaglandin D2 receptor PTGDR Q13258

Prostaglandin E2 receptor EP 1 subtype PTGER1 P34995

Prostaglandin E2 receptor EP2 subtype PTGER2 P431 16

Prostaglandin E2 receptor EP3 subtype PTGER3 P431 15

Prostaglandin E2 receptor EP4 subtype PTGER4 P35408

Type-2 angiotensin II receptor (AT2) AGTR2 P50052

Apelin receptor APLNR P35414

B 1 bradykinin receptor BDKRB 1 P46663

B2 bradykinin receptor BDKRB2 P30411

C5a anaphylatoxin chemotactic receptor C5AR1 P21730

Cholecystokinin receptor type A CCKAR P32238

C-C chemokine receptor type 10 CCR10 P46092

C-C chemokine receptor type 1 CCR1 P32246

C-C chemokine receptor type 2 CCR2 P41597

C-C chemokine receptor type 3 CCR3 P51677

C-C chemokine receptor type 4 CCR4 P51679

C-C chemokine receptor type 5 CCR5 P51681

C-C chemokine receptor type 6 CCR6 P51684

C-C chemokine receptor type 7 CCR7 P32248

C-C chemokine receptor type 8 CCR8 P51685

C-C chemokine receptor type 9 CCR9 P51686

Cysteinyl leukotriene receptor 1 CYSLTR1 Q9Y271

Cysteinyl leukotriene receptor 2 CYSLTR2 Q9NS75

Cannabinoid receptor 1 CNR1 P21554

Cannabinoid receptor 2 CNR2 P34972

CX3C chemokine receptor 1 CX3CR1 P49238

High affinity interleukin-8 receptor A IL8RA P25024

High affinity interleukin-8 receptor B IL8RB P25025

C-X-C chemokine receptor type 3 CXCR3 P49682

C-X-C chemokine receptor type 4 CXCR4 P61073

C-X-C chemokine receptor type 6 CXCR6 000574

C-X-C chemokine receptor type 7 CXCR7 P25106 D(1A) dopamine receptor DRD1 P21728

D(2) dopamine receptor DRD2 P14416

D(3) dopamine receptor DRD3 P35462

D(4) dopamine receptor DRD4 P21917

D(1B) dopamine receptor DRD5 P21918

Melanocyte-stimulating hormone receptor MC1R 001726

Melatonin receptor type 1A MTNR1A P48039

Melatonin receptor type IB MTNR1B P49286

Substance-P receptor TACR1 P25103

Substance-K receptor TACR2 P21452

Neuromedin-K receptor TACR3 P29371

Neuromedin-B receptor NMBR P28336

Neuropeptides B/W receptor type 1 NPBWR1 P48145

Neuropeptides B/W receptor type 2 NPBWR2 P48146

Neuropeptide FF receptor 1 NPFFR1 Q9GZQ6

Neuropeptide FF receptor 2 NPFFR2 Q9Y5X5

Neuropeptide Y receptor type 1 NPY1R P25929

Neuropeptide Y receptor type 2 NPY2R P49146

Neuropeptide Y receptor type 4 PPYR1 P50391

Neuropeptide Y receptor type 5 NPY5R Q15761

Neurotensin receptor type 1 NTSR1 P30989

Neurotensin receptor type 2 NTSR2 095665

Olfactory receptor 10A2 OR10A2 Q9H208

Olfactory receptor 10A3 OR10A3 P58181

Olfactory receptor 10A4 OR10A4 Q9H209

Olfactory receptor 10A5 OR10A5 Q9H207

Olfactory receptor 10A6 OR10A6 Q8NH74

Olfactory receptor 10A7 OR10A7 Q8NGE5

Olfactory receptor 10AD1 OR10AD 1 Q8NGE0

Olfactory receptor 10AG1 OR10AG1 Q8NH19

Olfactory receptor 10C1 OR10C1 Q96KK4

Olfactory receptor 10G2 OR10G2 Q8NGC3

Olfactory receptor 10G3 OR10G3 Q8NGC4

Olfactory receptor 10G4 OR10G4 Q8NGN3

Olfactory receptor 10G6 OR10G6 Q8NH81

Olfactory receptor 10G7 OR10G7 Q8NGN6

Olfactory receptor 10G8 OR10G8 Q8NGN5

Olfactory receptor 2T10 OR2T10 Q8NGZ9

Olfactory receptor 2T11 OR2T11 Q8NH01

Olfactory receptor 2T12 OR2T12 Q8NG77

Olfactory receptor 2T27 OR2T27 Q8NH04

Olfactory receptor 2T29 OR2T29 Q8NH02

Olfactory receptor 2T33 OR2T33 Q8NG76

Olfactory receptor 2T34 OR2T34 Q8NGX1

Olfactory receptor 2T35 OR2T35 Q8NGX2

Olfactory receptor 4A15 OR4A15 Q8NGL6

Olfactory receptor 4A16 OR4A16 Q8NH70

Olfactory receptor 4A47 OR4A47 Q6IF82 Olfactory receptor 4C45 OR4C45 A6NMZ5

Olfactory receptor 4C46 OR4C46 A6NHA9

Olfactory receptor 4F15 OR4F15 Q8NGB8

Olfactory receptor 4F17 OR4F17 Q8NGA8

Olfactory receptor 4F21 OR4F21 095013

Olfactory receptor 51 A2 OR51A2 Q8NGJ7

Olfactory receptor 51 A4 OR51A4 Q8NGJ6

Olfactory receptor 51 A7 OR51A7 Q8NH64

Olfactory receptor 51 B2 OR51B2 Q9Y5P1

Olfactory receptor 51 B4 OR51B4 Q9Y5P0

Olfactory receptor 51 B5 OR51B5 Q9H339

Olfactory receptor 51 B5 OR51B6 Q9H340

Olfactory receptor 5 ID 1 OR51D1 Q8NGF3

Olfactory receptor 5 IE 1 OR51E1 Q8TCB6

Olfactory receptor 51 E2 OR51E2 Q9H255

Olfactory receptor 5 IF 1 OR51F1 A6NGY5

Olfactory receptor 51 F2 OR51F2 Q8NH61

Olfactory receptor 51G1 OR51G1 Q8NGK1

Olfactory receptor 51 G2 OR51G2 Q8NGK0

Putative olfactory receptor 51H1 OR51H1P Q8NH63

Olfactory receptor 5111 OR51I1 Q9H343

Olfactory receptor 56A1 OR56A1 Q8NGH5

Olfactory receptor 56A3 OR56A3 Q8NH54

Olfactory receptor 56A4 OR56A4 Q8NGH8

Olfactory receptor 56A5 OR56A5 P0C7T3

Olfactory receptor 56B 1 OR56B 1 Q8NGI3

Olfactory receptor 56B4 OR56B4 Q8NH76

Olfactory receptor 5AC2 OR5AC2 Q9NZP5

Olfactory receptor 5AK2 OR5AK2 Q8NH90

Olfactory receptor 5 AN 1 OR5AN1 Q8NGI8

Olfactory receptor 5AP2 OR5AP2 Q8NGF4

Olfactory receptor 5AR1 OR5AR1 Q8NGP9

Olfactory receptor 5AS1 OR5AS 1 Q8N127

Olfactory receptor 5AU1 OR5AU1 Q8NGC0

Olfactory receptor 5H14 OR5H14 A6NHG9

Olfactory receptor 5H15 OR5H15 A6NDH6

Olfactory receptor 6C65 OR6C65 A6NJZ3

Olfactory receptor 6C68 OR6C68 A6NDL8

Olfactory receptor 6C70 OR6C70 A6NIJ9

Olfactory receptor 6C74 OR6C74 A6NCV1

Olfactory receptor 6C75 OR6C75 A6NL08

Olfactory receptor 6C76 OR6C76 A6NM76

Olfactory receptor 7E24 OR7E24 Q6IFN5

Opsin-3 OPN3 Q9H1Y3

Melanopsin OPN4 Q9UHM6

Opsin-5 OPN5 Q6U736 δ-type opioid receptor OPRD1 P41143

K-type opioid receptor OPRK1 P41145 μ-type opioid receptor OPRM1 P35372

Nociceptin receptor OPRL1 P41146

Blue-sensitive opsin OPN1SW P03999

Rhodopsin RHO P08100

Green-sensitive opsin OPN1MW P04001

Olfactory receptor 2B 11 OR2B11 Q5JQS5

Olfactory receptor 2C 1 OR2C1 095371

Olfactory receptor 2C3 OR2C3 Q8N628

Olfactory receptor 2D2 OR2D2 Q9H210

Olfactory receptor 2D 3 OR2D3 Q8NGH3

Olfactory receptor 2F 1 OR2F1 Q13607

Olfactory receptor 2F2 OR2F2 095006

Olfactory receptor 2G2 OR2G2 Q8NGZ5

Olfactory receptor 2G3 OR2G3 Q8NGZ4

Olfactory receptor 2G6 OR2G6 Q5TZ20

Olfactory receptor 2H1 OR2H1 Q9GZK4

Olfactory receptor 2H2 OR2H2 095918

Putative olfactory receptor 211 OR2I1P Q8NGU4

Olfactory receptor 2J1 OR2J1 Q9GZK6

Olfactory receptor 2J2 OR2J2 076002

Olfactory receptor 2J3 OR2J3 076001

Olfactory receptor 2K2 OR2K2 Q8NGT1

Olfactory receptor 2L2 OR2L2 Q8NH16

Olfactory receptor 2L3 OR2L3 Q8NG85

Olfactory receptor 2L5 OR2L5 Q8NG80

Olfactory receptor 2L8 OR2L8 Q8NGY9

Olfactory receptor 2L13 OR2L13 Q8N349

Olfactory receptor 2M2 OR2M2 Q96R28

Olfactory receptor 2M3 OR2M3 Q8NG83

Olfactory receptor 2M4 OR2M4 Q96R27

Olfactory receptor 2M5 OR2M5 A3KFT3

Olfactory receptor 2M7 OR2M7 Q8NG81

Olfactory receptor 2S2 OR2S2 Q9NQN1

Olfactory receptor 2T1 OR2T1 043869

Olfactory receptor 2T2 OR2T2 Q6IF00

Olfactory receptor 2T3 OR2T3 Q8NH03

Olfactory receptor 2T4 OR2T4 Q8NH00

Olfactory receptor 4E1 OR4E1 P0C645

Olfactory receptor 4E2 OR4E2 Q8NGC2

Olfactory receptor 4F3/4F16/4F29 OR4F3 Q6IEY1

Olfactory receptor 4F4 OR4F4 Q96R69

Olfactory receptor 4F5 OR4F5 Q8NH21

Olfactory receptor 4F6 OR4F6 Q8NGB9

Olfactory receptor 4K1 OR4K1 Q8NGD4

Olfactory receptor 4K2 OR4K2 Q8NGD2

Olfactory receptor 4K3 OR4K3 Q96R72

Olfactory receptor 4K5 OR4K5 Q8NGD3

Olfactory receptor 4K13 OR4K13 Q8NH42 Olfactory receptor 4K14 OR4K14 Q8NGD5

Olfactory receptor 4K15 OR4K15 Q8NH41

Olfactory receptor 4K17 OR4K17 Q8NGC6

Olfactory receptor 4L1 OR4L1 Q8NH43

Olfactory receptor 4M1 OR4M1 Q8NGD0

Olfactory receptor 4M2 OR4M2 Q8NGB6

Olfactory receptor 4N2 OR4N2 Q8NGD 1

Olfactory receptor 4N4 OR4N4 Q8N0Y3

Olfactory receptor 4N5 OR4N5 Q8IXE1

Olfactory receptor 4P4 OR4P4 Q8NGL7

Olfactory receptor 4Q2 OR4Q2 P0C623

Olfactory receptor 4Q3 OR4Q3 Q8NH05

Olfactory receptor 4S 1 OR4S1 Q8NGB4

Olfactory receptor 4S2 OR4S2 Q8NH73

Olfactory receptor 4X 1 OR4X1 Q8NH49

Olfactory receptor 4X2 OR4X2 Q8NGF9

Olfactory receptor 5 A 1 OR5A1 Q8NGJ0

Olfactory receptor 5A2 OR5A2 Q8NGI9

Olfactory receptor 5B2 OR5B2 Q96R09

Olfactory receptor 5B3 OR5B3 Q8NH48

Olfactory receptor 5B 12 OR5B12 Q96R08

Olfactory receptor 6A2 OR6A2 095222

Olfactory receptor 6B 1 OR6B1 095007

Olfactory receptor 6B2 OR6B2 Q6IFH4

Olfactory receptor 6B3 OR6B3 Q8NGW1

Olfactory receptor 6C 1 OR6C1 Q96RD1

Olfactory receptor 6C2 OR6C2 Q9NZP2

Olfactory receptor 6C3 OR6C3 Q9NZP0

Olfactory receptor 6C4 OR6C4 Q8NGE1

Olfactory receptor 6C6 OR6C6 A6NF89

Olfactory receptor 6F 1 OR6F1 Q8NGZ6

Olfactory receptor 6J1 OR6J1 Q8NGC5

Olfactory receptor 6K2 OR6K2 Q8NGY2

Olfactory receptor 6K3 OR6K3 Q8NGY3

Olfactory receptor 6K6 OR6K6 Q8NGW6

Olfactory receptor 6M1 OR6M1 Q8NGM8

Olfactory receptor 6N 1 OR6N1 Q8NGY5

Olfactory receptor 6N2 OR6N2 Q8NGY6

Olfactory receptor 6P 1 OR6P1 Q8NGX9

Olfactory receptor 6Q 1 OR6Q1 Q8NGQ2

Olfactory receptor 6S 1 OR6S1 Q8NH40

Olfactory receptor 6T1 OR6T1 Q8NGN1

Olfactory receptor 6V 1 OR6V1 Q8N148

Olfactory receptor 6X 1 OR6X1 Q8NH79

Olfactory receptor 6Y 1 OR6Y1 Q8NGX8

Olfactory receptor 7A5 OR7A5 015622

Olfactory receptor 7A10 OR7A10 076100

Olfactory receptor 7A17 OR7A17 014581 Olfactory receptor 7C 1 OR7C1 076099

Olfactory receptor 7C2 OR7C2 060412

Olfactory receptor 7D4 OR7D4 Q8NG98

Olfactory receptor 7G1 OR7G1 Q8NGA0

Olfactory receptor 7G2 OR7G2 Q8NG99

Prostaglandin F2-a receptor PTGFR P43088

Prostacyclin receptor PTGIR P431 19

Prolactin-releasing peptide receptor PRLHR P49683

Platelet-activating factor receptor PTAFR P25105

Pyroglutamylated RFamide peptide receptor QRFPR Q96P65

RPE-retinal G protein-coupled receptor RGR P47804

Sphingosine 1 -phosphate receptor 1 S 1PR1 P21453

Sphingosine 1 -phosphate receptor 2 S 1PR2 095136

Sphingosine 1 -phosphate receptor 3 S 1PR3 099500

Sphingosine 1 -phosphate receptor 4 S 1PR4 095977

Sphingosine 1 -phosphate receptor 5 S 1PR5 Q9H228

Somatostatin receptor type 1 SSTR1 P30872

Somatostatin receptor type 2 SSTR2 P30874

Somatostatin receptor type 3 SSTR3 P32745

Somatostatin receptor type 4 SSTR4 P31391

Somatostatin receptor type 5 SSTR5 P35346

Thromboxane A2 receptor TBXA2R P21731

Trace amine-associated receptor 1 TAAR1 Q96RJ0

Trace amine-associated receptor 2 TAAR2 Q9P1P5

Putative trace amine-associated receptor 3 TAAR3 Q9P1P4

Trace amine-associated receptor 5 TAAR5 014804

Trace amine-associated receptor 6 TAAR6 Q96RI8

Trace amine-associated receptor 8 TAAR8 Q969N4

Trace amine-associated receptor 9 TAAR9 Q96RI9

Thyrotropin receptor TSHR PI 6473

Vasopressin Via receptor AVPR1A P37288

Vasopressin Vlb receptor AVPR1B P47901

Vasopressin V2 receptor AVPR2 P30518

Chemokine XC receptor 1 XCR1 P46094

Brain-specific angiogenesis inhibitor 1 BAI1 014514

Brain-specific angiogenesis inhibitor 2 BAI2 060241

Brain-specific angiogenesis inhibitor 3 BAI3 060242

Calcitonin receptor CALCR P30988

Calcitonin gene-related peptide type 1 receptor CALCRL 016602

Corticotropin-releasing factor receptor 1 CRHR1 P34998

Corticotropin-releasing factor receptor 2 CRHR2 Q13324

Growth hormone-releasing hormone receptor GHRHR Q02643

Gastric inhibitory polypeptide receptor GIPR P48546

Glucagon-like peptide 1 receptor GLP1R P43220

Glucagon-like peptide 2 receptor GLP2R 095838

Glucagon receptor GCGR P47871

Pituitary adenylate cyclase-activating ADCYAP1R1 P41586 polypeptide type I receptor Taste receptor 1 member 2 TAS 1R2 Q8TE23

Parathyroid hormone receptor 1 PTH1R 003431

Parathyroid hormone 2 receptor PTH2R P49190

Secretin receptor SCTR P47872

Vasoactive intestinal polypeptide receptor 1 VIPR1 P32241

Vasoactive intestinal polypeptide receptor 2 VIPR2 P41587

Frizzled- 10 FZD 10 Q9ULW2

Frizzled- 1 FZD 1 Q9UP38

Frizzled-2 FZD2 014332

Frizzled-3 FZD3 Q9NPG1

Frizzled-4 FZD4 Q9ULV1

Frizzled-5 FZD5 Q 13467

Frizzled-6 FZD6 060353

Frizzled-7 FZD7 075084

Frizzled- 8 FZD8 Q9H461

Frizzled-9 FZD9 (FZD3) 000144

Smoothened homologue SMO (SMOH) Q99835

Extracellular calcium-sensing receptor CASR P41180

GAB A type B receptor lsubunit 1 GABBR1 Q9UBS5

GABA type B receptor subunit 2 GABBR2 075899

GPCR family C group 6 member A GPRC6A Q5T6X5

Metabotropic glutamate receptor 1 GRM1 Q 13255

Metabotropic glutamate receptor 2 GRM2 Q 14416

Metabotropic glutamate receptor 3 GRM3 Q 14832

Metabotropic glutamate receptor 4 GRM4 Q 14833

Metabotropic glutamate receptor 5 GRM5 P41594

Metabotropic glutamate receptor 6 GRM6 015303

Metabotropic glutamate receptor 7 GRM7 Q 14831

Metabotropic glutamate receptor 8 GRM8 000222

Taste receptor 1 member 1 TAS 1R1 Q7RTX 1

Taste receptor 1 member 2 TAS 1R2 Q8TE23

Taste receptor 1 member 3 TAS 1R3 Q7RTX0

[0065] Described herein are GPCR binding domains, wherein the GPCR binding domains are designed based on surface interactions between a GPCR ligand and the GPCR. In some instances, the ligand is a subatomic particle (e.g. , a photon), an ion, an organic molecule, a peptide, and a protein. Non-limiting examples of ligands which can be bound by a GPCR include (-)-adrenaline, (-)-noradrenaline, (lyso)phospholipid mediators, [des-Arg lO]kallidin, [des-Arg9]bradykinin, [des-Glnl4]ghrelin, [Hyp3]bradykinin, [Leu] enkephalin,

[Met]enkephalin, 12-hydroxyheptadecatrienoic acid, 12R-HETE, 12S-HETE, 12S-HPETE, 15S- HETE, 17 -estradiol, 20-hydroxy-LTB4, 2-arachidonoylglycerol, 2-oleoyl-LPA, 3- hydroxyoctanoic acid, 5-hydroxytryptamine, 5-oxo- 15-HETE, 5-oxo-ETE, 5-oxo-ETrE, 5-oxo- ODE, 5S-HETE, 5S-HPETE, 7a,25-dihydroxy cholesterol, acetylcholine, ACTH, adenosine diphosphate, adenosine, adrenomedullin 2/intermedin, adrenomedullin, amylin, anandamide, angiotensin II, angiotensin III, annexin I, apelin receptor early endogenous ligand, apelin-13, apelin-17, apelin-36, aspirin triggered lipoxin A4, aspirin-triggered resolvin Dl, ATP, beta- defensin 4A, big dynorphin, bovine adrenal medulla peptide 8-22, bradykinin, C3a, C5a, Ca2+, calcitonin gene related peptide, calcitonin, cathepsin G, CCK-33, CCK-4, CCK-8, CCL1, CCL11, CCL13, CCL14, CCL15, CCL16, CCL17, CCL19, CCL2, CCL20, CCL21, CCL22, CCL23, CCL24, CCL25, CCL26, CCL27, CCL28, CCL3, CCL4, CCL5, CCL7, CCL8, chemerin, chenodeoxycholic acid, cholic acid, corticotrophin-releasing hormone, CST-17, CX3CL1, CXCL1, CXCL10, CXCL11, CXCL12a, ΟΧ(Χ12β, CXCL13, CXCL16, CXCL2, CXCL3, CXCL5, CXCL6, CXCL7, CXCL8, CXCL9, cysteinyl-leukotrienes (CysLTs), uracil nucleotides, deoxycholic acid, dihydrosphingosine-1 -phosphate, dioleoylphosphatidic acid, dopamine, dynorphin A, dynorphin A-(l-13), dynorphin A-(l-8), dynorphin B, endomoφhin-l, endothelin-1, endothelin-2, endothelin-3, F2L, Free fatty acids, FSH, GABA, galanin, galanin- like peptide, gastric inhibitory polypeptide, gastrin- 17, gastrin-releasing peptide, ghrelin, GHRH, glucagon, glucagon-like peptide l-(7-36) amide, glucagon-like peptide l-(7-37), glucagon-like peptide 2, glucagon-like peptide 2-(3-33), GnRH I, GnRH II, GRP-( 18-27), hCG, histamine, humanin, INSL3, INSL5, kallidin, kisspeptin-10, kisspeptin-13, kisspeptin-14, kisspeptin-54, kynurenic acid, large neuromedin N, large neurotensin, L-glutamic acid, LH, lithocholic acid, L- lactic acid, long chain carboxylic acids, LPA, LTB4, LTC4, LTD4, LTE4, LXA4, Lys-[Hyp3]- bradykinin, lysophosphatidylinositol, lysophosphatidylserine, Medium-chain-length fatty acids, melanin-concentrating hormone, melatonin, methylcarbamyl PAF, Mg2+, motilin, N- arachidonoylglycine, neurokinin A, neurokinin B, neuromedin B, neuromedin N, neuromedin S- 33, neuromedin U-25, neuronostatin, neuropeptide AF, neuropeptide B-23, neuropeptide B-29, neuropeptide FF, neuropeptide S, neuropeptide SF, neuropeptide W-23, neuropeptide W-30, neuropeptide Y, neuropeptide Y-(3-36), neurotensin, nociceptin/oφhanin FQ, N- oleoylethanolamide, obestatin, octopamine, orexin-A, orexin-B, Oxysterols, oxytocin, PACAP- 27, PACAP-38, PAF, pancreatic polypeptide, peptide YY, PGD2, PGE2, PGF2a, PGI2, PGJ2, PHM, phosphatidylserine, PHV, prokineticin-1, prokineticin-2, prokineticin-2 , prosaposin, PrRP-20, PrRP-31, PTH, PTHrP, PTHrP-(l-36), QRFP43, relaxin, relaxin-1, relaxin-3, resolvin Dl, resolvin El, RFRP-1, RFRP-3, R-spondins, secretin, serine proteases, sphingosine 1- phosphate, sphingosylphosphorylcholine, SRIF-14, SRIF-28, substance P, succinic acid, thrombin, thromboxane A2, TIP39, T-kinin, TRH, TSH, tyramine, UDP-glucose, uridine diphosphate, urocortin 1, urocortin 2, urocortin 3, urotensin II -related peptide, urotensin-II, vasopressin, VIP, Wnt, Wnt-1, Wnt-lOa, Wnt-lOb, Wnt-11, Wnt-16, Wnt-2, Wnt-2b, Wnt-3, Wnt-3a, Wnt-4, Wnt-5a, Wnt-5b, Wnt-6, Wnt-7a, Wnt-7b, Wnt-8a, Wnt-8b, Wnt-9a, Wnt-9b, XCL1, XCL2, Zn2+, a-CGRP, a-ketoglutaric acid, a-MSH, a-neoendoφhin, β-alanine, β- CGPvP, β-D-hydroxybutyric acid, β-endoφhin, β-MSH, β-neoendoφhin, β-phenylethylamine, and γ-MSH.

[0066] Sequences of GPCR binding domains based on surface interactions between a GPCR ligand and the GPCR are analyzed using various methods. For example, multispecies computational analysis is performed. In some instances, a structure analysis is performed. In some instances, a sequence analysis is performed. Sequence analysis can be performed using a database known in the art. Non-limiting examples of databases include, but are not limited to, NCBI BLAST (blast.ncbi.nlm.nih.gov/Blast.cgi), UCSC Genome Browser (genome.ucsc.edu/), UniProt (www.uniprot.org/), and IUPHAR BPS Guide to PHARMACOLOGY

(guidetopharmacology . org/) .

[0067] Described herein are GPCR binding domains designed based on sequence analysis among various organisms. For example, sequence analysis is performed to identify homologous sequences in different organisms. Exemplary organisms include, but are not limited to, mouse, rat, equine, sheep, cow, primate (e.g., chimpanzee, baboon, gorilla, orangutan, monkey), dog, cat, pig, donkey, rabbit, fish, fly, and human.

[0068] Following identification of GPCR binding domains, libraries comprising nucleic acids encoding for the GPCR binding domains may be generated. In some instances, libraries of GPCR binding domains comprise sequences of GPCR binding domains designed based on conformational ligand interactions, peptide ligand interactions, small molecule ligand interactions, extracellular domains of GPCRs, or antibodies that target GPCRs. Libraries of GPCR binding domains may be translated to generate protein libraries. In some instances, libraries of GPCR binding domains are translated to generate peptide libraries, immunoglobulin libraries, derivatives thereof, or combinations thereof. In some instances, libraries of GPCR binding domains are translated to generate protein libraries that are further modified to generate peptidomimetic libraries. In some instances, libraries of GPCR binding domains are translated to generate protein libraries that are used to generate small molecules.

[0069] Methods described herein provide for synthesis of libraries of GPCR binding domains comprising nucleic acids each encoding for a predetermined variant of at least one predetermined reference nucleic acid sequence. In some cases, the predetermined reference sequence is a nucleic acid sequence encoding for a protein, and the variant library comprises sequences encoding for variation of at least a single codon such that a plurality of different variants of a single residue in the subsequent protein encoded by the synthesized nucleic acid are generated by standard translation processes. In some instances, the libraries of GPCR binding domains comprise varied nucleic acids collectively encoding variations at multiple positions. In some instances, the variant library comprises sequences encoding for variation of at least a single codon in a GPCR binding domain. In some instances, the variant library comprises sequences encoding for variation of multiple codons in a GPCR binding domain. An exemplary number of codons for variation include, but are not limited to, at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 225, 250, 275, 300, or more than 300 codons.

[0070] Methods described herein provide for synthesis of libraries comprising nucleic acids encoding for the GPCR binding domains, wherein the libraries comprise sequences encoding for variation of length of the GPCR binding domains. In some instances, the library comprises sequences encoding for variation of length of at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 225, 250, 275, 300, or more than 300 codons less as compared to a predetermined reference sequence. In some instances, the library comprises sequences encoding for variation of length of at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 275, 300, or more than 300 codons more as compared to a predetermined reference sequence.

[0071] Following identification of GPCR binding domains, the GPCR binding domains may be placed in scaffolds as described herein. In some instances, the scaffolds are

immunoglobulins. In some instances, the GPCR binding domains are placed in the CDR-H3 region. GPCR binding domains that may be placed in scaffolds can also be referred to as a motif. Scaffolds comprising GPCR binding domains may be designed based on binding, specificity, stability, expression, folding, or downstream activity. In some instances, the scaffolds comprising GPCR binding domains enable contact with the GPCRs. In some instances, the scaffolds comprising GPCR binding domains enables high affinity binding with the GPCRs. Exemplary amino acid sequences of GPCR binding domains are described in Table 2.

Table 2. GPCR amino acid sequences

ALKTTVILILAFFACWLPYYIGISIDSFILLEIIKQGCEFENTVHKWISIT

EALAFFHCCLNPILYAFLGAKFKTSAQHALTSVSRGSSLKILSKGKR

GGHSSVSTESESSSFHSS

CCR4 MNPTDIADTTLDESIYSNYYLYESIPKPCTKEGIKAFGELFLPPLYSL

VFVFGLLGNSVWLVLFKYKRLRSMTDVYLLNLAISDLLFVFSLPF

WGYYAADQWVFGLGLCKMISWMYLVGFYSGIFFVMLMSIDRYLAI

VHAVFSLRARTLTYGVITSLATWSVAVFASLPGFLFSTCYTERNHTY

CKTKYSLNSTTWKVLSSLEINILGLVIPLGIMLFCYSMIIRTLQHCKN

EKKNKAVKMIFAVWLFLGFWTPYNIVLFLETLVELEVLQDCTFER

YLDYAIQATETLAFVHCCLNPIIYFFLGEKFRKYILQLFKTCRGLFVL

CQYCGLLQIYSADTPSSSYTQSTMDHDLHDAL

GCGR MPPCQPQRPLLLLLLLLACQPQVPSAQVMDFLFEKWKLYGDQCHH

NLSLLPPPTELVCNRTFDKYSCWPDTPANTTANISCPWYLPWHHKV

QHRFVFKRCGPDGQWVRGPRGQPWRDASQCQMDGEEIEVQKEVA

KMYSSFQVMYTVGYSLSLGALLLALAILGGLSKLHCTRNAIHANLF

ASFVLKASSVLVIDGLLRTRYSQKIGDDLSVSTWLSDGAVAGCRVA

AVFMQYGIVANYCWLLVEGLYLHNLLGLATLPERSFFSLYLGIGWG

APMLFVVPWAVVKCLFENVQCWTSNDNMGFWWILRFPVFLAILIN

FFIFVRIVQLLVAKLRARQMHHTDYKFRLAKSTLTLIPLLGVHEVVF

AFVTDEHAQGTLRSAKLFFDLFLSSFQGLLVAVLYCFLNKEVQSEL

RRRWHRWRLGKVLWEERNTSNHRASSSPGHGPPSKELQFGRGGGS

QDSSAETPLAGGLPRLAESPF

mGluR5 MVLLLILSVLLLKEDVRGSAQSSERRVVAHMPGDIIIGALFSVHHQP

TVDKVHERKCGAVREQYGIQRVEAMLHTLERINSDPTLLPNITLGC

EIRD SCWHS A V ALEQ SIEFIRD SLI S SEEEEGLVRCVDGS S S SFRSKKP

IVGVIGPGSSSVAIQVQNLLQLFNIPQIAYSATSMDLSDKTLFKYFMR

VVPSDAQQARAMVDIVKRYNWTYVSAVHTEGNYGESGMEAFKD

MSAKEGICIAHSYKIYSNAGEQSFDKLLKKLTSHLPKARVVACFCEG

MTVRGLLMAMRRLGLAGEFLLLGSDGWADRYDVTDGYQREAVG

GmKLQSPDVKWFDDYYLKLRPETNHRNPWFQEFWQHRFQCRLEG

FPQENSKYNKTCNSSLTLKTHHVQDSKMGFVINAIYSMAYGLHNM

QMSLCPGYAGLCDAMKPIDGRKLLESLMKTNFTGVSGDTILFDENG

DSPGRYEIMNFKEMGKDYFDYINVGSWDNGELKMDDDEVWSKKS

NIIRSVCSEPCEKGQIKVIRKGEVSCCWTCTPCKENEYVFDEYTCKA

CQLGSWPTDDLTGCDLIPVQYLRWGDPEPIAAVVFACLGLLATLFV

TVVFIIYRDTPVVKSSSRELCYIILAGICLGYLCTFCLIAKPKQIYCYL

QRIGIGLSPAMSYSALVTKTNRIARILAGSKKKICTKKPRFMSACAQ

LVIAFILICIQLGIIVALFIMEPPDIMHDYPSIREVYLICNTTNLGWTP

LGYNGLLILSCTFYAFKTRNVPANFNEAKYIAFTMYTTCIIWLAFVPI

YFGSNYKIITMCFSVSLSATVALGCMFVPKVYIILAKPERNVRSAFTT

STVVRMHVGDGKSSSAASRSSSLVNLWKRRGSSGETLRYKDRRLA

QHKSEIECFTPKGSMGNGGRATMSSSNGKSVTWAQNEKSSRGQHL

WQRLSIHINKKENPNQTAVIKPFPKSTESRGLGAGAGAGGSAGGVG

ATGGAGCAGAGPGGPESPDAGPKALYDVAEAEEHFPAPARPRSPSP

ISTLSHRAGSASRTDDDVPSLHSEPVARSSSSQGSLMEQISSVVTRFT

ANISELNSMMLSTAAPSPGVGAPLCSSYLIPKEIQLPTTMTTFAEIQP

LPAIEVTGGAQPAAGAQAAGDAARESPAAGPEAAAAKPDLEELVA

LTPPSPFRDSVDSGSTTPNSPVSESALCIPSSPKYDTLIIRDYTQSSSSL

GLP-1R RPQGATVSLWETVQKWREYRRQCQRSLTEDPPPATDLFCNRTFDE

YACWPDGEPGSFVNVSCPWYLPWASSVPQGHVYRFCTAEGLWLQ KDNSSLPWRDLSECEESKRGERSSPEEQLLFLYIIYTVGYALSFSALV IASAILLGFRHLHCTO YIHLNLFASFILRALSVFIKDAALKWMYSTA

AQQHQWDGLLSYQDSLSCRLVFLLMQYCVAANYYWLLVEGVYLY

TLLAFSVLSEQWIFRLYVSIGWGVPLLFVVPWGIVKYLYEDEGCWT

RNSNMNYWLIIRLPILFAIGVNFLIFVRVICIVVSKLKANLMCKTDIK

CRLAKSTLTLIPLLGTHEVIFAFVMDEHARGTLRFIKLFTELSFTSFQ

GLMVAILYCFVN EVQLEFRKSWERWRLEHLHIQRDSSMKPLKCPT

SSLSSGATAGSSMYTATCQASCS

GABAB MLLLLLLAPLFLRPPGAGGAQTPNATSEGCQIIHPPWEGGIRYRGLT

RDQVKAINFLPVDYEIEYVCRGEREVVGPKVRKCLANGSWTDMDT

PSRCVRICSKSYLTLENGKVFLTGGDLPALDGARVDFRCDPDFHLV

GSSRSICSQGQWSTPKPHCQVNRTPHSERRAVYIGALFPMSGGWPG

GQACQPAVEMALEDVNSRRDILPDYELKLIHHDSKCDPGQATKYL

YELLYNDPIKIILMPGCSSVSTLVAEAARMWNLIVLSYGSSSPALSN

RQRFPTFFRTHPSATLHNPTRVKLFEKWGWKKIATIQQTTEVFTSTL

DDLEERVKEAGIEITFRQSFFSDPAVPVKNLKRQDARIIVGLFYETEA

RKVFCEVYKERLFGKKYVWFLIGWYADNWFKIYDPSINCTVDEMT

EAVEGHITTEIVMLNPANTRSISNMTSQEFVEKLTKRLKRHPEETGG

FQEAPLAYDAIWALALALNKTSGGGGRSGVRLEDFNYN QTITDQI

YRAMNSSSFEGVSGHVVFDASGSRMAWTLIEQLQGGSYKKIGYYD

STKDDLSWSKTDKWIGGSPPADQTLVIKTFRFLSQKLFISVSVLSSLG

IVLAVVCLSFNIYNSHVRYIQNSQPNLN LTAVGCSLALAAVFPLGL

DGYHIGRNQFPFVCQARLWLLGLGFSLGYGSMFTKIWWVHTVFTK

KEEKKEWRKTLEPWKLYATVGLLVGMDVLTLAIWQIVDPLHRTIE

TFAKEEPKEDIDVSILPQLEHCSSRKMNTWLGIFYGYKGLLLLLGIFL

AYETKSVSTEKINDHRAVGMAIYNVAVLCLITAPVTMILSSQQDAA

FAFASLAIVFSSYITLVVLFVPKMRRLITRGEWQSEAQDTMKTGSST

N EEEKSRLLEKENRELEKIIAEKEERVSELRHQLQSRQQLRSRRH

PPTPPEPSGGLPRGPPEPPDRLSCDGSRVHLLYK

OPRM1 MDSSAAPTNASNCTDALAYSSCSPAPSPGSWVNLSHLDGNLSDPCG

PNRTDLGGRDSLCPPTGSPSMITAITIMALYSIVCVVGLFGNFLVMY

VIVRYTKMKTATNIYIFNLALADALATSTLPFQSVNYLMGTWPFGΉ

LCKIVISIDYYNMFTSIFTLCTMSVDRYIAVCHPVKALDFRTPRNAKI

INVCNWILSSAIGLPVMFMATTKYRQGSIDCTLTFSHPTWYWENLL

KICVFIFAFIMPVLIITVCYGLMILRLKSVRMLSGSKEKDRNLRRITR

MVLVVVAVFIVCWTPIHIYVIIKALVTIPETTFQTVSWHFCIALGYTN

SCLNPVLYAFLDENFKRCFREFCIPTSSNIEQQNSTRIRQNTRDHPST

ANTVDRTNHQLENLEAETAPLP

OPRK1 MD SPIQIFRGEPGPTCAP S ACLPPN S S AWFPGWAEPD SNGS AGSED A

QLEPAfflSPAIPVIITAVYSVVFVVGLVGNSLVMFVIIRYTKMKTATN r^IFNLALADALVTTTMPFQSTVYLMNSWPFGDVLCKIVISIDYYNM

FTSIFTLTMMSVDRYIAVCHPVKALDFRTPLKAKIINICIWLLSSSVGI

SAIVLGGTKVREDVDVIECSLQFPDDDYSWWDLFMKICVFIFAFVIP

VLIIIVCYTLMILRLKSVRLLSGSREKDRNLRRITRLVLVVVAVFVVC

WTPIHIFILVEALGSTSHSTAALSSYYFCIALGYTNSSLNPILYAFLDE

NFKRCFRDFCFPLKMRMERQSTSRVRNTVQDPAYLRDIDGMNKPV

C5aR MDSFNYTTPDYGHYDDKDTLDLNTPVDKTSNTLRVPDILALVIFAV

VFLVGVLGNALVVWVTAFEAKRΉNAIWFLNLAVADFLSCLALPIL

FTSIVQHHHWPFGGAACSILPSLILLNMYASILLLATISADRFLLVFKP

IWCQNFRGAGLAWIACAVAWGLALLLTIPSFLYRVVREEYFPPKVL

CGVDYSHDKRRERAVAIVRLVLGFLWPLLTLTICYTFILLRTWSRRA

TRSTKTLKVVVAVVASFFIFWLPYQVTGIMMSFLEPSSPTFLLLKKL DSLCVSFAYINCCINPIIYVVAGQGFQGRLRKSLPSLLRNVLTEESVV RESKSFTRSTVDTMAQKTQAV

CGRP ELEESPEDSIQLGVTRNKIMTAQYECYQKIMQDPIQQAEGVYCNRT

WDGWLCWNDVAAGTESMQLCPDYFQDFDPSEKVTKICDQDGNWF

RHPASNRTWTNYTQCNVNTHEKVKTALNLFYLTIIGHGLSIASLLIS

LGIFFΎFKSLSCQRITLHKNLFFSFVCNSVVΉIHLTAVAN QALVAT

NPVSCKVSQFIHLYLMGCNYFWMLCEGIYLHTLIVVAVFAEKQHL

MWYYFLGWGFPLIPACIHAIARSLYYNDNCWISSDTHLLYIIHGPIC

AALLVNLFFLLNIVRVLITKLKVTHQAESNLYMKAVRATLILVPLLG

IEFVLIPWRPEGKIAEEVYDYIMHILMHFQGLLVSTIFCFFNGEVQAI

LRRNWNQYKIQFGNSFSNSEALRSASY ^Γ ΓVSΉSDGPGYSHDCPSEHL

NGKSIHDIENVLLKPENLYN

Ml MNTSAPPAVSPNITVLAPGKGPWQVAFIGITTGLLSLATVTGNLLVL muscarinic ISFKVNTELK N YFLLSLACADLIIGTFSMNLYTTYLLMGHWAL

GTLACDLWLALDYVASNASVMNLLLISFDRYFSVTRPLSYRAKRTP

RRAALMIGLAWLVSFVLWAPAILFWQYLVGERTVLAGQCYIQFLS

QPIITFGTAMAAFYLPVTVMCTLYWRIYRETENRARELAALQGSETP

GKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEE

EEDEGSMESLTSSEGEEPGSEVVIKMPMVDPEAQAPTKQPPRSSPNT

VKRPTKKGRDRAGKGQKPRGKEQLAKRKTFSLVKEKKAARTLSAI

LLAFILTWTPYNIMVLVSTFCKDCVPETLWELGYWLCYVNSTINPM

CYALCNKAFRDTFRLLLLCRWDKRRWRKIPKRPGSVHRTPSRQC

M4 MANFTPVNGSSGNQSVRLVTSSSHNRYETVEMVFIATVTGSLSLVT muscarinic VVGNILVMLSIKVNRQLQ N YFLFSLACADLIIGAFSMNLYTVYI

IKGYWPLGAVVCDLWLALDYVVSNASVMNLLIISFDRYFCVTKPLT

YPARRTTKMAGLMIAAAWVLSFVLWAPAILFWQFVVGKRTVPDN

QCFIQFLSNPAVTFGTAIAAFYLPVVIMTVLYIHISLASRSRVHKHRP

EGPKEKKAKTLAFLKSPLMKQSVKKPPPGEAAREELRNGKLEEAPP

PALPPPPRPVADKDTSNESSSGSATQNTKERPATELSTTEATTPAMP

APPLQPRALNPASRWSKIQIVTKQTGNECVTAIEIVPATPAGMRPAA

NVARKFASIARNQVRKKRQMAARERKVTRTIFAILLAFILTWTPYN

VMVLVNTFCQSCIPDTVWSIGYWLCYVNSTINPACYALCNATFKKT

FRHLLLCQYRNIGTAR

CCR2 MLSTSRSRFIRNTNESGEEVTTFFDYDYGAPCHKFDVKQIGAQLLPP

LYSLVFIFGFVGNMLVVLILINCKKLKCLTDIYLLNLAISDLLFLITLP

LWAHSAANEWVFGNAMCKLFTGLYHIGYFGGIFFIILLTIDRYLAIV

HAVFALKARTVTFGVVTSVITWLVAVFASVPGIIFTKCQKEDSVYV

CGPYFPRGWN FHTIMRNILGLVLPLLIMVICYSGILKTLLRCRNEK

KRHRAVRVIFΉMIVYFLFWTPYNIVILLNTFQEFFGLSNCESTSQLD

QATQVTETLGMTHCCINPIIYAFVGEKFRSLFHIALGCRIAPLQKPVC

GGPGVRPGKNVKVTTQGLLDGRGKGKSIGRAPEASLQDKEGA

CCR9 MTPTDFTSPIPNMADDYGSESTSSMEDYVNFNFTDFYCEKN VRQF

ASHFLPPLYWLVFIVGALGNSLVILVYWYCTRVKTMTDMFLLNLAI

ADLLFLVTLPFWAIAAADQWKFQTFMCKVVNSMYKMNFYSCVLLI

MCISVDRYIAIAQAMRAHTWREKRLLYSKMVCFTIWVLAAALCIPE

ILYSQIKEESGIAICTMVYPSDESTKLKSAVLTLKVILGFFLPFVVMA

CCYTfflHTLIQAKKSSKHKALKVTI L FVLSQFPYNCILLV(^D

AYAMFISNCAVSTNIDICFQVTQTIAFFHSCLNPVLYVFVGERFRRDL

VKTLKNLGCISQAQWVSFTRREGSLKLSSMLLETTSGALSL

GPR174 MPANYTCTRPDGDNTDFRYFIYAVTYTVILVPGLIGNILALWVFYG

YMKETKRAVIFMINLAIADLLQVLSLPLRIFYYLNHDWPFGPGLCMF CFYLKYVNMYASIYFLVCISVRRFWFLMYPFRFHDCKQKYDLYISIA

GWLIICLACVLFPLLRTSDDTSGNRTKCFVDLPTRNVNLAQSVVMM

TIGELIGFVTPLLIVLYCTWKTVLSLQDKYPMAQDLGEKQKALKMI

LTCAGVFLICFAPYHFSFPLDFLVKSNEIKSCLARRVILIFHSVALCLA

SLNSCLDPVIYYFSTNEFRRRLSRQDLHDSIQLHAKSFVSNHTASTM

TPELC

MASP-2 TPLGPKWPEPVFGRLASPGFPGEYANDQERRWTLTAPPGYRLRLYF

THFDLELSHLCEYDFVKLSSGAKVLATLCGQESTDTERAPGKDTFY

SLGSSLDITFRSDYSNEKPFTGFEAFYAAEDIDECQVAPGEAPTCDH

HCHNHLGGFYCSCRAGYVLHRNKRTCSALCSGQVFTQRSGELSSPE

YPRPYPKLSSCTYSISLEEGFSVILDFVESFDVETHPETLCPYDFLKIQ

TDREEHGPFCGKTLPHRIETKSNTVTITFVTDESGDHTGWKIHYTST

AQPCPYPMAPPNGHVSPVQAKYILKDSFSIFCETGYELLQGHLPLKS

FTAVCQKDGSWDRPMPACSIVDCGPPDDLPSGRVEYITGPGVTTYK

AVIQYSCEETFYTMKVNDGKYVCEADGFWTSSKGEKSLPVCEPVC

GLSARTTGGRIYGGQKAKPGDFPWQVLILGGTTAAGALLYDNWVL

TAAHAVYEQKHDASALDIRMGTLKRLSPHYTQAWSEAVFIHEGYT

HDAGFDNDIALIKLN KVVINSNITPICLPRKEAESFMRTDDIGTASG

WGLTQRGFLARNLMYVDIPIVDHQKCTAAYEKPPYPRGSVTANML

CAGLESGGKDSCRGDSGGALVFLDSETERWFVGGIVSWGSMNCGE

AGQYGVYTKVINYIPWIENIISDF

CCR5 MDYQVSSPIYDINYYTSEPCQKINVKQIAARLLPPLYSLVFIFGFVGN

MLVILILINCKRLKSMTDIYLLNLAISDLFFLLTVPFWAHYAAAQWD

FGNTMCQLLTGLYFIGFFSGIFFIILLTIDRYLAVVHAVFALKARTVT

FGVVTSVITWVVAVFASLPGIIFTRSQKEGLHYTCSSHFPYSQYQFW

KNFQTLKIVILGLVLPLLVMVICYSGILKTLLRCRNEKKRHRAVRLIF

TIMIVYFLFWAPYNIVLLLNTFQEFFGLN CSSSNRLDQAMQVTETL

GMTHCCINPIIYAFVGEKFRNYLLVFFQKHIAKRFCKCCSIFQQEAPE

RA S S VYTRSTGEQEIS VGL

FSHR CHHRICHCSNRVFLCQESKVTEIPSDLPRNAIELRFVLTKLRVIQKGA

FSGFGDLEKIEISQNDVLEVIEADVFSNLPKLHEIRIEKAN LLYINPE

AFQNLPNLQYLLISNTGIKHLPDVHKIHSLQKVLLDIQDNINIHTIER

NSFVGLSFESVILWLNKNGIQEIHNCAFNGTQLDELNLSDN LEEL

PNDVFHGASGPVILDISRTRIHSLPSYGLENLKKLRARSTYNLKKLPT

LEKLVALMEASLTYPSHCCAFANWRRQISELHPICNKSILRQEVDY

MTQARGQRSSLAEDNESSYSRGFDMTYTEFDYDLCNEVVDVTCSP

KPDAFNPCEDIMGYNILRVLIWFISILAITGNIIVLVILTTSQYKLTVPR

FLMCNLAFADLCIGIYLLLIASVDIHTKSQYHNYAIDWQTGAGCDA

AGFF FASELS TLTAITLERWHTITHAMQLDCKVQLRHAASVM

VMGWIFAFAAALFPIFGISSYMKVSICLPMDIDSPLSQLYVMSLLVL

NVLAF VVICGCYIHrfLTVRNPNIV S S S SDTRIAKRMAMLIFTDFLCM

APISFFAISASLKVPLITVSKAKILLVLFHPINSCANPFLYAIFTKNFRR

DFFILLSKCGCYEMQAQIYRTETSSTVHNTHPRNGHCSSAPRVTNGS

TYILVPLSHLAQN

mGluR2 EGPAKKVLTLEGDLVLGGLFPVHQKGGPAEDCGPVNEHRGIQRLEA PAM MLFALDRINRDPHLLPGVRLGAHILDSCSKDTHALEQALDFVRASLS

RGADGSRHICPDGSYATHGDAPTAITGVIGGSYSDVSIQVANLLRLF

QIPQISYASTSAKLSDKSRYDYFARTVPPDFFQAKAMAEILRFFNWT

YVSTVASEGDYGETGIEAFELEARARNICVATSEKVGRAMSRAAFE

GVVRALLQKPSARVAVLFTRSEDARELLAASQRLNASFTWVASDG

WGALESVVAGSEGAAEGAITIELASYPISDFASYFQSLDPWNNSRNP WFREFWEQRFRCSFRQRDCAAHSLRAVPFEQESKIMFVVNAVYAM

AHALHNMHRALCPNTTRLCDAMRPVNGRRLYKDFVLNVKFDAPF

RPADTHNEVRFDRFGDGIGRYNIFTYLRAGSGRYRYQKVGYWAEG

LTLDTSLIPWASPSAGPLPASRCSEPCLQNEVKSVQPGEVCCWLCIP

CQPYEYRLDEFTCADCGLGYWPNASLTGCFELPQEYIRWGDAWAV

GPVTIACLGALATLFVLGVFVRHNATPVVKASGRELCYILLGGVFL

CYCMTFIFIAKPSTAVCTLRRLGLGTAFSVCYSALLTKTNRIARIFGG

AREGAQRPRFISPASQVAICLALISGQLLIVVAWLVVEAPGTGKETA

PERREVVTLRCNHRDASMLGSLAYNVLLIALCTLYAFKTRKCPENF

NEAKFIGFTMYTTCIIWLAFLPIFYVTSSDYRVQTTTMCVSVSLSGSV

VLGCLFAPKLHIILFQPQKNVVSHRAPTSRFGSAAARASSSLGQGSG

S QFVPTVCNGREVVD STTS SL

mGluR3 LGDHNFLRREIKIEGDLVLGGLFPINEKGTGTEECGRINEDRGIQRLE

AMLFAIDEINKDDYLLPGVKLGVHILDTCSRDTYALEQSLEFVRASL

TKVDEAEYMCPDGSYAIQENIPLLIAGVIGGSYSSVSIQVANLLRLFQ

IPQISYASTSAKLSDKSRYDYFARTVPPDFYQAKAMAEILRFFNWTY

VSTVASEGDYGETGIEAFEQEARLRNICIATAEKVGRSNIRKSYDSVI

RELLQKPNARWVLFMRSDDSRELIAAASRANASFTWVASDGWGA

QESIIKGSEHVAYGAITLELASQPVRQFDRYFQSLNPYN HRNPWFR

DFWEQKFQCSLQNKRNHRRVCDKHLAIDSSNYEQESKIMFVVNAV

YAMAHALHKMQRTLCPNTTKLCDAMKILDGKKLYKDYLLKINFTA

PFNPNKDADSIVKFDTFGDGMGRYNVFNFQNVGGKYSYLKVGHW

AETLSLDVNSIHWSRNSVPTSQCSDPCAPNEMKNMQPGDVCCWICI

PCEPYEYLADEFTCMDCGSGQWPTADLTGCYDLPEDYIRWEDAWA

IGPVTIACLGFMCTCMVVTVFIKHN TPLVKASGRELCYILLFGVGL

SYCMTFFFIAKPSPVICALRRLGLGSSFAICYSALLTKTNCIARIFDGV

KNGAQRPKFISPSSQVFICLGLILVQIVMVSVWLILEAPGTRRYTLAE

KRETVILKCNVKDSSMLISLTYDVILVILCTVYAFKTRKCPENFNEA

KFIGFTMYTTCIIWLAFLPIFYVTSSDYRVQTTTMCISVSLSGFVVLG

CLFAPKVHIILFQPQKNVVTHRLHLNRFSVSGTGTTYSQSSASTYVP

TVCNGREVLDSTTSSL

mGluR4 KPKGHPHMNSIRIDGDITLGGLFPVHGRGSEGKPCGELKKEKGIHRL

EAMLFALDRIN DPDLLPNITLGARILDTC SRDTHALEQ SLTFVQ ALI

EKDGTEVRCGSGGPPIITKPERVVGVIGASGSSVSIMVANILRLFKIP

QISYASTAPDLSDNSRYDFFSRVVPSDTYQAQAMVDIVRALKWNY

VSTVASEGSYGESGVEAFIQKSREDGGVCIAQSVKIPREPKAGEFDKI

IRRLLETSNARAVIIFANEDDIRRVLEAARRANQTGHFFWMGSDSW

GSKIAPVLHLEEVAEGAVTILPKRMSVRGFDRYFSSRTLDNNRRNIW

FAEFWEDNFHCKLSRHALKKGSHVKKCTNRERIGQDSAYEQEGKV

QFVIDAVYAMGHALHAMHRDLCPGRVGLCPRMDPVDGTQLLKYI

RNVNFSGIAGNPVTFNENGDAPGRYDIYQYQLRNDSAEYKVIGSWT

DHLHLRIERMHWPGSGQQLPRSICSLPCQPGERKKTVKGMPCCWH

CEPCTGYQYQVDRYTCKTCPYDMRPTENRTGCRPIPIIKLEWGSPW

AVLPLFLAVVGIAATLFWITFVRYNDTPIVKASGRELSYVLLAGIFL

CYATTFLMIAEPDLGTCSLRRIFLGLGMSISYAALLTKTNRIYRIFEQ

GKRSVSAPRFISPASQLAITFSLISLQLLGICVWFVVDPSHSVVDFQD

QRTLDPRFARGVLKCDISDLSLICLLGYSMLLMVTCTVYAIKTRGVP

ETFNEAKPIGFTMYTTCIVWLAFIPIFFGTSQSADKLYIQTTTLTVSVS

LSASVSLGMLYMPKVYIILFHPEQNVPKRKRSLKAVVTAATMSNKF

TQKGNFRPNGEAKSELCENLEAPALATKQTYVTYTNHAI

mGluR7 QEMYAPHSIRIEGDVTLGGLFPVHAKGPSGVPCGDIKRENGIHRLEA MLYALDQINSDPNLLPNVTLGARILDTCSRDTYALEQSLTFVQALIQ

KDTSDVRCTNGEPPVFVKPEKVVGVIGASGSSVSIMVANILRLFQIP

QISYASTAPELSDDRRYDFFSRVVPPDSFQAQAMVDIVKALGWNYV

STLASEGSYGEKGVESFTQISKEAGGLCIAQSVRIPQERKDRTIDFDR

IIKQLLDTPNSRAVVIFANDEDIKQILAAAKRADQVGHFLWVGSDS

WGSKINPLHQHEDIAEGAITIQPKRATVEGFDAYFTSRTLEN RRNV

\νΡΑΕΥ\νΕΕΝΡΝ0ΚΕΉ8Ο8ΚΚΕΟΤΟ^€ΤθρΕ� �ΟΚΟ8ΝΥΕρΕΟΚ

VQFVIDAVYAMAHALHHMNKDLCADYRGVCPEMEQAGGKKLLK

YIRNVNFNGSAGTPVMFNKNGDAPGRYDIFQYQTTNTSNPGYRLIG

QWTDELQLNIEDMQWGKGVREIPASVCTLPCKPGQRKKTQKGTPC

CWTCEPCDGYQYQFDEMTCQHCPYDQRPNENRTGCQDIPIIKLEWH

SPWAVIPVFLAMLGIIATIFVMATFIRYNDTPIVRASGRELSYVLLTGI

FLCYIITFLMIAKPDVAVCSFRRVFLGLGMCISYAALLTKTNRIYRIF

EQGKKSVTAPRLISPTSQLAITSSLISVQLLGVFIWFGVDPPNIIIDYDE

HKTMNPEQARGVLKCDITDLQIICSLGYSILLMVTCTVYAIKTRGVP

ENFNEAKPIGFTMYTTCI VWLAFIPIFFGTAQ S AEKLYIQTTTLTI SMN

LSASVALGMLYMPKVYIIIFHPELNVQKRKRSFKAVVTAATMSSRL

SHKPSDRPNGEAKTELCENVDPNSPAAKKKYVSYN LVI

CXCR3 MVLEVSDHQVLNDAEVAALLENFSSSYDYGENESDSCCTSPPCPQD

FSLNFDRAFLPALYSLLFLLGLLGNGAVAAVLLSRRTALSSTDTFLL

HLAVADTLLVLTLPLWAVDAAVQWVFGSGLCKVAGALFNINFYA

GALLLACISFDRYLNIVHATQLYRRGPPARVTLTCLAVWGLCLLFA

LPDFIFLSAHHDERLNATHCQYNFPQVGRTALRVLQLVAGFLLPLL

VMAYCYAfflLAVLLVSRGQRRLRAMRLVVVVVVAFALCWTPYHL

VVLVDILMDLGALARNCGRESRVDVAKSVTSGLGYMHCCLNPLLY

AFVGVKFRERMWMLLLRLGCPNQRGLQRQPSSSRRDSSWSETSEA

SYSGL

CCR8 MDYTLDLSVTTVTDYYYPDIFSSPCDAELIQTNGKLLLAVFYCLLFV

FSLLGNSLVILVLVVCKKLRSITDVYLLNLALSDLLFVFSFPFQTYYL

LDQWVFGTVMCKVVSGFYYIGFYSSMFFITLMSVDRYLAVVHAVY

ALKVRΉRMGTTLCLAVWLTAIMATIPLLVFΎQVASEDGVLQCYSF

YNQQTLKWKIFTNFKMNILGLLIPFTIFMFCYIKILHQLKRCQNHNK

TKAIRLVLIVVIASLLFWVPFNVVLFLTSLHSMHILDGCSISQQLTYA

TWTEnSFTHCCVNPVIYAFVGEKFKKHLSEIFQKSCSQIFNYLGRQ

MPRESCEKSSSCQQHSSRSSSVDYIL

Adenosine MPIMGSSVYITVELAIAVLAILGNVLVCWAVWLNSNLQNVTNYFV A2a VSLAAADIAVGVLAIPFAITISTGFCAACHGCLFIACFVLVLTQSSIFS

LLAIAIDRYIAIRIPLRYNGLVTGTRAKGIIAICWVLSFAIGLTPMLGW

N CGQPKEGKNHSQGCGEGQVACLFEDVVPMNYMVYFNFFACVL

VPLLLMLGVYLRIFLAARRQLKQMESQPLPGERARSTLQKEVHAAK

SLAIIVGLFALCWLPLHIINCFTFFCPDCSHAPLWLMYLAIVLSHTNS

VVNPFIYAYRIREFRQTFRKIIRSHVLRQQEPFKAAGTSARVLAAHG

SDGEQVSLRLNGHPPGVWANGSAPHPERRPNGYALGLVSGGSAQE

SQGNTGLPDVELLSHELKGVCPEPPGLDDPLAQDGAGVS

Orexin MEPSATPGAQMGVPPGSREPSPVPPDYEDEFLRYLWRDYLYPKQYE 0X1 W LIAAYVAVFVVALVGNTLVCLAVWRNHHMRTVTNYFIVNLSL

ADVLVTAICLPASLLVDITESWLFGHALCKVIPYLQAVSVSVAVLTL

SFIALDRWYAICHPLLFKSTARRARGSILGIWAVSLAIMVPQAAVME

CSSVLPELANRTRLFSVCDERWADDLYPKIYHSCFFIVTYLAPLGLM

AMAYFQIFRKLWGRQIPGTTSALVRNWKRPSDQLGDLEQGLSGEPQ

PRARAFLAEVKQMRARRKTAKMLMVVLLVFALCYLPISVLNVLKR VFGMFRQASDREAVYACFTFSHWLVYANSAANPIIYNFLSGKFREQ

FKAAFSCCLPGLGPCGSLKAPSPRSSASHKSLSLQSRCSISKISEHVVL

TSVTTVLP

Orexin MSGTKLEDSPPCRNWSSASELNETQEPFLNPTDYDDEEFLRYLWRE 0X2 YLHPKEYEW LIAGYnVFVVALIGNVLVCVAVWKNHHMRTVTNY

FIVNLSLADVLVTITCLPATLVVDITETWFFGQSLCKVIPYLQTVSVS

VSVLTLSCIALDRWYAICHPLMFKSTAKRARNSIVIIWIVSCIIMIPQA

IVMECS FPGLANKTTLF CDERWGGErV KMYHICFFLVTYMA

PLCLMVLAYLQIFRKLWCRQIPGTSSVVQRKWKPLQPVSQPRGPGQ

PTKSRMSAVAAEIKQIRARRKTARMLMIVLLVFAICYLPISILNVLKR

VFGMFAHTEDRE YAWFTFSHWLVYANSAANPIIYNFLSGKFREE

FKAAFSCCCLGVHHRQEDRLTRGRTSTESRKSLTTQISNFDNISKLSE

Q VVLTSI STLP AANGAGPLQNW

PAR-2 IQGTNRSSKGRSLIGKVDGTSHVTGKGVTVETVFSVDEFSASVLTGK

LT FLPI TIVFVVGLPSNGMALWVFLFRTKKKHPAVIYMANLA

LADLLSVIWFPLKIAYHIHGN WIYGEALCNVLIGFFYGNMYCSILF

MTCLSVQRYWVIVNPMGHSRKKANIAIGISLAIWLLILLVTIPLYVV

KQTIFIPALNITTCHDVLPEQLLVGDMFNYFLSLAIGVFLFPAFLTAS

AYVLMIRMLRSSAMDENSEKKRKRAIKLIVTVLAMYLICFTPSNLLL

VVHYFLIKSQGQSHVYALYIVALCLSTLNSCIDPFVYYFVSHDFRDH

AKNALLCRSVRTVKQMQVSLTSKKHSRKS S SYSS SSTTVKTSY

C3aR MASFSAETOSTDLLSQPWNEPPVILSMVILSLTFLLGLPGNGLVLWV

AGLKMQRTVNTIWFLHLTLADLLCCLSLPFSLAHLALQGQWPYGRF

LCKLIPSIIVLNMFASVFLLTAISLDRCLVVFKPIWCQNHRNVGMAC

SICGCIWVVAFVMCIPVFVYREIFTTDNHNRCGYKFGLSSSLDYPDF

YGDPLENRSLENIVQPPGEMNDRLDPSSFQTNDHPWTVPTVFQPQT

FQRPSADSLPRGSARLTSQNLYSNVFKPADVVSPKIPSGFPIEDHETS

PLDNSDAFLSTHLKLFPSASSNSFYESELPQGFQDYYNLGQFTDDDQ

VPTPLVAITITRLVVGFLLPSVIMIACYSFIVFRMQRGRFAKSQSKTF

RVAVVVVAVFLVCWTPYHIFGVLSLLTDPETPLGKTLMSWDHVCIA

LASANSCFNPFLYALLGKDFRKKARQSIQGILEAAFSEELTRSTHCPS

N VISERNSTTV

LGR5 GSSPRSGVLLRGCPTHCHCEPDGRMLLRVDCSDLGLSELPSNLSVFT

SYLDLSMN ISQLLPNPLPSLRFLEELRLAGNALTYIPKGAFTGLYSL

KVLMLQN QLRHVPTEALQNLRSLQSLRLDANHISYVPPSCFSGLH

SLRHLWLDDNALTEIPVQAFRSLSALQAMTLALNKIHHIPDYAFGN

LSSLVVLHLHN RIHSLGKKCFDGLHSLETLDLNYN LDEFPTAIRT

LSNLKELGFHSN IRSIPEKAFVGNPSLITIHFYDNPIQFVGRSAFQHL

PELRTLTLNGASQITEFPDLTGTANLESLTLTGAQISSLPQTVCNQLP

NLQVLDLSYNLLEDLPSFSVCQKLQKIDLRHNEIYEIKVDTFQQLLS

LRSLNLAWNKIAIIHPNAFSTLPSLIKLDLSSNLLSSFPITGLHGLTHL

KLTGNHALQSLISSENFPELKVIEMPYAYQCCAFGVCENAYKISNQ

WNKGDNSSMDDLHKKDAGMFQAQDERDLEDFLLDFEEDLKALHS

VQCSPSPGPFKPCEHLLDGWLIRIGVWΉAVLALTCNALVTS ^Γ ΓVFRS

PLYISPIKLLIGVIAAVNMLTGVSSAVLAGVDAFTFGSFARHGAWW

ENGVGCHVIGFLSIFASESSVFLLTLAALERGFSVKYSAKFETKAPFS

SLKVIILLCALLALTMAAVPLLGGSKYGASPLCLPLPFGEPSTMGYM

VALILLNSLCFLMMTIAYTKLYCNLDKGDLENIWDCSMVKHIALLL

FTNCILNCPVAFLSFSSLINLTFISPEVIKFILLVVVPLPACLNPLLYILF

NPHFKEDLVSLRKQTYVWTRSKHPSLMSINSDDVEKQSCDSTQALV

TFTSSSITYDLPPSSVPSPAYPVTESCHLSSVAFVPCL GPR101 MTSTCTNSTRESNSSHTCMPLSKMPISLAHGIIRSTVLVIFLAASFVG

NIVLALVLQRKPQLLQVTNRFIFNLLVTDLLQISLVAPWVVATSVPL

FWPLNSHFCTALVSLTHLFAFASVNTIVVVSVDRYLSIIHPLSYPSKM

TQRRGYLLLYGTWIVAILQSTPPLYGWGQAAFDERNALCSMIWGA

SPSYTILSVVSFIVIPLIVMIACYSVVFCAARRQHALLYNVKRHSLEV

RVKDCVENEDEEGAEKKEEFQDESEFRRQHEGEVKAKEGRMEAKD

GSLKAKEGSTGTSESSVEARGSEEVRESSTVASDGSMEGKEGSTKV

EENSMKADKGRTEVNQCSIDLGEDDMEFGEDDINFSEDDVEAVNIP

ESLPPSRRNSNSNPPLPRCYQCKAAKVIFIIIFSYVLSLGPYCFLAVLA

VW DVETQVPQWVITIIIWLFFLQCCIHPYVYGYMHKTIKKEIQDM

LKKFFCKEKPPKEDSHPDLPGTEGGTEGKIVPSYDSATFP

GPR151 MLAAAFAD SNS S SMNVSF AHLHFAGGYLP SD S QDWRTIIPALLVA V

CLVGFVGNLCVIGILLHNAWKGKPSMIHSLILNLSLADLSLLLFSAPI

RATAYSKSVWDLGWFVCKSSDWFIHTCMAAKSLTIVVVAKVCFM

YASDPAKQVSIHNYTIWSVLVAIWTVASLLPLPEWFFSTIRHHEGVE

MCLVDVPAVAEEFMSMFGKLYPLLAFGLPLFFASFYFWRAYDQCK

KRGTKTQNLRNQIRSKQV MLLSIAnSALLWLPEWVAWLWVWH

LKAAGPAPPQGFIALSQVLMFSISSANPLIFLVMSEEFREGLKGVWK

WMITKKPPTVSESQETPAGNSEGLPDKVPSPESPASIPEKEKPSSPSS

GKGKTEKAEIPILPDVEQFWHERDTVPSVQDNDPIPWEHEDQETGE

GVK

GPR161 MSLNSSLSCRKELSNLTEEEGGEGGVIITQFIAIIVITIFVCLGNLVIW

TLYKKSYLLTLSNKFVFSLTLSNFLLSVLVLPFVVTSSIRREWIFGVV

WCNFSALLYLLISSASMLTLGVIAIDRYYAVLYPMVYPMKITGNRA

VMALVYIWLHSLIGCLPPLFGWSSVEFDEFKWMCVAAWHREPGYT

AFWQIWCALFPFLVMLVCYGFIFRVARVKARKVHCGTVVIVEEDA

QRTGRKNSSTSTSSSGSRRNAFQGVVYSANQCKALITILVVLGAFM

VTWGPYMVVIASEALWGKSSVSPSLETWATWLSFASAVCHPLIYGL

WNKTVRKELLGMCFGDRYYREPFVQRQRTSRLFSISNRITDLGLSPH

LTALMAGGQPLGHSSSTGDTGFSCSQDSGTDMMLLEDYTSDDNPPS

HCTCPPKRRSSVTFEDEVEQIKEAAKNSILHVKAEVHKSLDSYAASL

AKAIEAEAKINLFGEEALPGVLVTARTVPGGGFGGRRGSRTLVSQR

LQLQSIEEGDVLAAEQR

GPR17 MSKRSWWAGSRKPPREMLKLSGSDSSQSMNGLEVAPPGLITNFSLA

TAEQCGQETPLENMLFASFYLLDFILALVGNTLALWLFIRDHKSGTP

ANVFLMHLAVADLSCVLVLPTRLVYHFSGNHWPFGEIACRLTGFLF

YLNMYASIYFLTCISADRFLAIVHPVKSLKLRRPLYAHLACAFLWV

VVAVAMAPLLVSPQTVQTNHTVVCLQLYREKASHHALVSLAVAFT

FPFITTVTCYLLIIRSLRQGLRVEKRLKTKAVRMIAIVLAIFLVCFVPY

HVNRSVYVLHYRSHGASCATQRILALANRITSCLTSLNGALDPIMYF

FVAEKFRHALCNLLCGKRLKGPPPSFEGKTNESSLSAKSEL

GPR183 MDIQMAN FTPPSATPQGNDCDLYAHHSTARIVMPLHYSLVFIIGLV

GNLLALVVIVQNRKKINSTTLYSTNLVISDILFTTALPTRIAYYAMGF

DWRIGDALCRITALVFYINTYAGVNFMTCLSIDRFIAVVHPLRYNKI

KRIEHAKGVCIFVWILVFAQTLPLLINPMSKQEAERITCMEYPNFEET

KSLPWILLGACFIGYVLPLIIILICYSQICCKLFRTAKQNPLTEKSGVN

KKALNTIILIIVVFVLCFTPYHVAIIQHMIKKLRFSNFLECSQRHSFQIS

LHFWCLMNFNCCMDPFIYFFACKGYKRKVMRMLKRQVSVSISSA

VKS APEENSREMTETQMMIHSKS SNGK

CRTH2 MSANATLKPLCPILEQMSRLQSHSNTSIRYIDHAAVLLHGLASLLGL

VENGVILFVVGCRMRQTVVTTWVLHLALSDLLASASLPFFTYFLAV GHSWELGTTFCKLHSSIFFLNMFASGFLLSAISLDRCLQVVRPVWAQ

NHRTVAAAHKVCLVLWALAVLNTVPYFVFRDTISRLDGRIMCYYN

VLLLNPGPDRDATCNSRQVALAVSKFLLAFLVPLAIIASSHAAVSLR

LQHRGRRRPGRFVRLVAAVVAAFALCWGPYHVFSLLEARAHANPG

LRPLVWRGLPFVTSLAFFNSVANPVLYVLTCPDMLRKLRRSLRTVL

ESVLVDDSELGGAGSSRRRRTSSTARSASPLALCSRPEEPRGPARLL

GWLLGSCAASPQTGPLNRALSSTSS

5-HT4 MDKLDANVSSEEGFGSVEKWLLTFLSTVILMAILGNLLVMVAVC

WDRQLRKIKTNYFIVSLAFADLLVSVLVMPFGAIELVQDIWIYGEVF

CLVRTSLDVLLTTASIFHLCCISLDRYYAICCQPLVYRNKMTPLRIAL

MLGGCW IPTFISFLPIMQGWN IGIIDLIEKRKFNQNSNSTYCVFM

VNKPYAITCSVVAFYIPFLLMVLAYYRIYVTAKEHAHQIQMLQRAG

ASSESRPQSADQHSTHRMRTETKAAKTLCIIMGCFCLCWAPFFVTNI

VDPFIDYTVPGQVWTAFLWLGYINSGLNPFLYAFLNKSFRRAFLIIL

CCDDERYRRPSILGQTVPCSTTTINGSTHVLRDAVECGGQWESQCH

PPATSPLVAAQPSDT

5-HT6 MVPEPGPTANSTPAWGAGPPSAPGGSGWVAAALCVVIALTAAANS

LLIALICTQPALRNTSNFFLVSLFTSDLMVGLVVMPPAMLNALYGR

WVLARGLCLLWTAFDVMCCSASILNLCLISLDRYLLILSPLRYKLR

MTPLRALALVLGAWSLAALASFLPLLLGWHELGHARPPVPGQCRL

LASLPFVLVASGLTFFLPSGAICFTYCRILLAARKQAVQVASLTTGM

ASQASETLQVPRTPRPGVESADSRRLATKHSRKALKASLTLGILLGM

FFVTWLPFFVANIVQAVCDCISPGLFDVLTWLGYCNSTMNPIIYPLF

MRDFKRALGRFLPCPRCPRERQASLASPSLRTSHSGPRPGLSLQQVL

PLPLPPD SD SD SD AGSGGS SGLRLTAQLLLPGEATQDPPLPTRAAAA

VNFFNIDPAEPELRPHPLGIPTN

CB2 MEECWVTEIANGSKDGLDSNPMKDYMILSGPQKTAVAVLCTLLGL

LSALENVAVLYLILSSHQLRRKPSYLFIGSLAGADFLASVVFACSFV

NFHVFHGVDSKAVFLLKIGSVTMTFTASVGSLLLTAIDRYLCLRYPP

SYKALLTRGRALVTLGIMWVLSALVSYLPLMGWTCCPRPCSELFPL

IPNDYLLSWLLFIAFLFSGIIYTYGHVLWKAHQHVASLSGHQDRQVP

GMARMRLDVRLAKTLGLVLAVLLICWFPVLALMAHSLATTLSDQV

KKAFAFCSMLCLINSMVNPVIYALRSGEIRSSAHHCLAHWKKCVRG

LGSEAKEEAPRSSVTETEADGKITPWPDSRDLDLSDC

Histamine- MERAPPDGPLNASGALAGEAAAAGGARGFSAAWTAVLAALMALL

3 IVATVLGNALVMLAFVADSSLRTQN FFLLNLAISDFLVGAFCIPLY

VPYVLTGRWTFGRGLCKLWLVVDYLLCTSSAFNIVLISYDRFLSVT

RAVSYRAQQGDTRRAVRKMLLVWVLAFLLYGPAILSWEYLSGGSS

IPEGHCYAEFFYNWYFLITASTLEFFTPFLSVTFFNLSIYLNIQRRTRL

RLDGAREAAGPEPPPEAQPSPPPPPGCWGCWQKGHGEAMPLHRYG

VGEAAVGAEAGEATLGGGGGGGSVASPTSSSGSSSRGTERPRSLKR

GSKPSASSASLEKRMKMVSQSFTQRFRLSRDRKVAKSLAVIVSIFGL

CWAPYTLLMIIRAACHGHCVPDYWYETSFWLLWANSAVNPVLYPL

CHHSFRRAFTKLLCPQKLKIQPHS SLEHCWK

VPAC-1 ARLQEECDYVQMIEVQHKQCLEEAQLENETIGCSKMWDNLTCWPA or VIPR1 TPRGQVVVLACPLIFKLFSSIQGRNVSRSCTDEGWTHLEPGPYPIAC

GLDDKAASLDEQQTMFYGSVKTGYTIGYGLSLATLLVATAILSLFR

KLHCTRNYIHMHLFISFILRAAAVFIKDLALFDSGESDQCSEGSVGC

KAAMVFFQYCVMANFFWLLVEGLYLYTLLAVSFFSERKYFWGYILI

Ο\νθνΡ8ΤΡΤΜν\νΤΙΑΡαΗΡΕΟΥΟ0\νθΉΝ88� �\ν\νΐΙΚΟΡΙΕΤ8ΙΕνΝ

FILFICIIRILLQKLRPPDIRKSDSSPYSRLARSTLLLIPLFGVHYIMFAF FPDNFKPEVKMVFELVVGSFQGFVVAILYCFLNGEVQAELRRKWR

RWHLQGVLGWNPKYRHPSGGSNGATCSTQVSMLTRVSPGARRSSS

FQAEVSLV

GIPR RAETGSKGQTAGELYQRWERYRRECQETLAAAEPPSGLACNGSFD

MYVCWDYAAPNATARASCPWYLPWHHHVAAGFVLRQCGSDGQW

GLWRDHTQCENPEKNEAFLDQRLILERLQVMYTVGYSLSLATLLLA

LLILSLFRRLHCTRNYIHINLFTSFMLRAAAILSRDRLLPRPGPYLGD

QALALWNQALAACRTAQIVTQYCVGANYTWLLVEGVYLHSLLVL

VGGSEEGHFRYYLLLGWGAPALFVIPWVIVRYLYENTQCWERNEV

KAIWWIIRTPILMTILINFLIFIRILGILLSKLRTRQMRCRDYRLRLARS

TLTLVPLLGVHEVVFAPVTEEQARGALRFAKLGFEIFLSSFQGFLVS

VLYCFINKEVQSEIRRGWHHCRLRRSLGEEQRQLPERAFRALPSGSG

PGEVPTSRGLSSGTLPGPGNEASRELESYC

5-HT1B MEEPGAQCAPPPPAGSETWVPQANLSSAPSQNCSAKDYIYQDSISLP GPCR WKVLLVMLLALITLATTLSNAFVIATVYRTRKLHTPANYLIASLAVT

DLLVSILVMPISTMYTVTGRWTLGQVVCDFWLSSDITCCTASILHLC

VIALDRYWAITDAVEYSAKRTPKRAAVMIALVWVFSISISLPPFFWR

QAKAEEEVSECVVNTDHILYTVYSTVGAFYFPTLLLIALYGRIYVEA

RSRILKQTPNRTGKRLTRAQLITDSPGSTSSVTSINSRVPDVPSESGSP

VYVNQVKVRVSDALLEKKKLMAARERKATKTLGIILGAFIVCWLPF

FIISLVMPICKDACWFHLAIFDFFTWLGYLNSLINPIIYTMSNEDFKQ

AFHKLIRFKCTS

CCR7 QDEVTDDYIGDNTTVDYTLFESLCSKKDVRNFKAWFLPIMYSIICFV

GLLGNGLVVLTYIYFKRLKTMTDTYLLNLAVADILFLLTLPFWAYS

AAKSWVFGVHFCKLIFAIYKMSFFSGMLLLLCISIDRYVAIVQAVSA

HRHRARVLLISKLSCVGIWILATVLSIPELLYSDLQRSSSEQAMRCSL

ITEHVEAFITIQVAQMVIGFLVPLLAMSFCYLVIIRTLLQARNFERNK

AIKVIIAVVVVFIVFQLPYNGVVLAQTVANFNITSSTCELSKQLNIAY

DVTYSLACVRCCVNPFLYAFIGVKFRNDLFKLFKDLGCLSQEQLRQ

CXCR5 MNYPLTLEMDLENLEDLFWELDRLDNYNDTSLVENHLCPATEGPL

MASFKAVFVPVAYSLIFLLGVIGNVLVLVILERHRQTRSSTETFLFHL

AVADLLLVFILPFAVAEGSVGWVLGTFLCKTVIALHKVNFYCSSLLL

ACIAVDRYLAIVHAVHAYRHRRLLSIHITCGTIWLVGFLLALPEILFA

KVSQGHHN SLPRCTFSQENQAETHAWFTSRFLYHVAGFLLPMLV

MGWCYVGVVHRLRQAQRRPQRQKAVRVAILVTSIFFLCWSPYHIVI

FLDTLARLKAVDNTCKLNGSLPVAITMCEFLGLAHCCLNPMLYTFA

GVKFRSDLSRLLTKLGCTGPASLCQLFPSWRRSSLSESENATSLTTF

GPR119 MESSFSFGVILAVLASLIIATNTLVAVAVLLLIHKNDGVSLCFTLNLA

VADTLIGVAISGLLTDQLSSPSRPTQKTLCSLRMAFVTSSAAASVLT

VMLITFDRYLAIKQPFRYLKIMSGFVAGACIAGLWLVSYLIGFLPLGI

PMFQQTAYKGQCSFFAVFHPHFVLTLSCVGFFPAMLLFVFFYCDML

KIASMHSQQIRKMEHAGAMAGGYRSPRTPSDFKALRTVSVLIGSFA

LSWTPFLITGIVQVACQECHLYLVLERYLWLLGVGNSLLNPLIYAY

WQKEVRLQLYHMALGVKKVLTSFLLFLSARNCGPERPRESSCHIVT

ISSSEFDG

GPR55 MSQQNTSGDCLFDGVNELMKTLQFAVHIPTFVLGLLLNLLAIHGFS

TFLKNRWPDYAATSIYMINLAVFDLLLVLSLPFKMVLSQVQSPFPSL

CTLVECLYFVSMYGSVFTICFISMDRFLAIRYPLLVSHLRSPRKIFGIC

CTIWVLVWTGSIPIYSFHGKVEKYMCFHNMSDDTWSAKVFFPLEVF

GFLLPMGIMGFCCSRSIHILLGRRDHTQDWVOOKACIYSIAASLAVF VVSFLPVHLGFFLQFLVRNSFIVECRAKQSISFFLQLSMCFSNVNCCL I DVFCYYFVIKEFRMNIRAHRPSRVQLVLQDTTISRG

[0072] Provided herein are scaffolds comprising GPCR binding domains, wherein the sequences of the GPCR binding domains support interaction with at least one GPCR. The sequence may be homologous or identical to a sequence of a GPCR ligand. In some instances, the GPCR binding domain sequence comprises at least or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,

32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47. In some instances, the GPCR binding domain sequence comprises at least or about 95% homology to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32,

33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47. In some instances, the GPCR binding domain sequence comprises at least or about 97% homology to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47. In some instances, the GPCR binding domain sequence comprises at least or about 99% homology to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47. In some instances, the GPCR binding domain sequence comprises at least or about 100% homology to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,

34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47. In some instances, the GPCR binding domain sequence comprises at least a portion having at least or about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, or more than 400 amino acids of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47.

[0073] Libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains may bind to one or more GPCRs. In some instances, the scaffolds comprising GPCR binding domains binds to a single GPCR. In some instances, the scaffolds comprising GPCR binding domains binds to GPCRs in a same family or class. In some instances, the scaffolds comprising GPCR binding domains bind to multiple GPCRs. For example, the scaffolds are multimeric and comprise at least 2 scaffolds. In some instances, the multimeric scaffolds comprise at least or about 3, 4, 5, 6, 7, 8, or more than 8 scaffolds. In some instances, the multimeric scaffolds comprise at least 2 scaffolds linked by, for example, a dimerization domain, an amino acid linker, a disulfide bond, a chemical crosslink, or any other linker known in the art. In some instances, the multimeric scaffolds bind to the same GPCRs or different GPCRs.

[0074] Provided herein are GPCR binding libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains comprise variation in domain type, domain length, or residue variation. In some instances, the domain is a region in the scaffold comprising the GPCR binding domains. For example, the region is the VH, CDR-H3, or VL domain. In some instances, the domain is the GPCR binding domain.

[0075] Methods described herein provide for synthesis of a GPCR binding library of nucleic acids each encoding for a predetermined variant of at least one predetermined reference nucleic acid sequence. In some cases, the predetermined reference sequence is a nucleic acid sequence encoding for a protein, and the variant library comprises sequences encoding for variation of at least a single codon such that a plurality of different variants of a single residue in the subsequent protein encoded by the synthesized nucleic acid are generated by standard translation processes. In some instances, the GPCR binding library comprises varied nucleic acids collectively encoding variations at multiple positions. In some instances, the variant library comprises sequences encoding for variation of at least a single codon of a VH, CDR-H3, or VL domain. In some instances, the variant library comprises sequences encoding for variation of at least a single codon in a GPCR binding domain. For example, at least one single codon of a GPCR binding domain as listed in Table 2 is varied. In some instances, the variant library comprises sequences encoding for variation of multiple codons of a VH, CDR-H3, or VL domain. In some instances, the variant library comprises sequences encoding for variation of multiple codons in a GPCR binding domain. An exemplary number of codons for variation include, but are not limited to, at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 225, 250, 275, 300, or more than 300 codons.

[0076] Methods described herein provide for synthesis of a GPCR binding library of nucleic acids each encoding for a predetermined variant of at least one predetermined reference nucleic acid sequence, wherein the GPCR binding library comprises sequences encoding for variation of length of a domain. In some instances, the domain is VH, CDR-H3, or VL domain. In some instances, the domain is the GPCR binding domain. In some instances, the library comprises sequences encoding for variation of length of at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 225, 250, 275, 300, or more than 300 codons less as compared to a predetermined reference sequence. In some instances, the library comprises sequences encoding for variation of length of at least or about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 275, 300, or more than 300 codons more as compared to a predetermined reference sequence.

[0077] Provided herein are GPCR binding libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains, wherein the GPCR binding libraries are synthesized with various numbers of fragments. In some instances, the fragments comprise the VH, CDR-H3, or VL domain. In some instances, the GPCR binding libraries are synthesized with at least or about 2 fragments, 3 fragments, 4 fragments, 5 fragments, or more than 5 fragments. The length of each of the nucleic acid fragments or average length of the nucleic acids synthesized may be at least or about 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, 600, or more than 600 base pairs. In some instances, the length is about 50 to 600, 75 to 575, 100 to 550, 125 to 525, 150 to 500, 175 to 475, 200 to 450, 225 to 425, 250 to 400, 275 to 375, or 300 to 350 base pairs.

[0078] GPCR binding libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains as described herein comprise various lengths of amino acids when translated. In some instances, the length of each of the amino acid fragments or average length of the amino acid synthesized may be at least or about 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, or more than 150 amino acids. In some instances, the length of the amino acid is about 15 to 150, 20 to 145, 25 to 140, 30 to 135, 35 to 130, 40 to 125, 45 to 120, 50 to 115, 55 to 110, 60 to 110, 65 to 105, 70 to 100, or 75 to 95 amino acids. In some instances, the length of the amino acid is about 22 to about 75 amino acids.

[0079] GPCR binding libraries comprising de novo synthesized variant sequences encoding for scaffolds comprising GPCR binding domains comprise a number of variant sequences. In some instances, a number of variant sequences is de novo synthesized for a CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, CDR-L3, VL, VH, or a combination thereof. In some instances, a number of variant sequences is de novo synthesized for framework element 1 (FWl), framework element 2 (FW2), framework element 3 (FW3), or framework element 4 (FW4). In some instances, a number of variant sequences is de novo synthesized for a GPCR binding domain. For example, the number of variant sequences is about 1 to about 10 sequences for the VH domain, about 10 ⁸ sequences for the GPCR binding domain, and about 1 to about 44 sequences for the VK domain. See FIG. 2. The number of variant sequences may be at least or about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, or more than 500 sequences. In some instances, the number of variant sequences is about 10 to 300, 25 to 275, 50 to 250, 75 to 225, 100 to 200, or 125 to 150 sequences.

[0080] GPCR binding libraries comprising de novo synthesized variant sequences encoding for scaffolds comprising GPCR binding domains comprise improved diversity. For example, variants are generated by placing GPCR binding domain variants in immunoglobulin scaffold variants comprising N-terminal CDR-H3 variations and C-terminal CDR-H3 variations. In some instances, variants include affinity maturation variants. Alternatively or in combination, variants include variants in other regions of the immunoglobulin including, but not limited to, CDR-H1, CDR-H2, CDR-L1, CDR-L2, and CDR-L3. In some instances, the number of variants of the GPCR binding libraries is least or about 10 ⁴ , 10 ⁵ , 10 ⁶ , 10 ⁷ , 10 ⁸ , 10 ⁹ , 10 ¹⁰ , or more than 10 ¹⁰ non- identical sequences. For example, a library comprising about 10 variant sequences for a VH region, about 237 variant sequences for a CDR-H3 region, and about 43 variant sequences for a VL and CDR-L3 region comprises 10 ⁵ non-identical sequences (10 x 237 x 43). See FIGS. 4A- 4B

[0081] Following synthesis of GPCR binding libraries comprising nucleic acids encoding scaffolds comprising GPCR binding domains, libraries may be used for screening and analysis. For example, libraries are assayed for library displayability and panning. In some instances, displayability is assayed using a selectable tag. Exemplary tags include, but are not limited to, a radioactive label, a fluorescent label, an enzyme, a chemiluminescent tag, a colorimetric tag, an affinity tag or other labels or tags that are known in the art. In some instances, the tag is histidine, polyhistidine, myc, hemagglutinin (HA), or FLAG. For example as seen in FIG. 3, the GPCR binding libraries comprises nucleic acids encoding scaffolds comprising GPCR binding domains with multiple tags such as GFP, FLAG, and Lucy as well as a DNA barcode. In some instances, libraries are assayed by sequencing using various methods including, but not limited to, single -molecule real-time (SMRT) sequencing, Polony sequencing, sequencing by ligation, reversible terminator sequencing, proton detection sequencing, ion semiconductor sequencing, nanopore sequencing, electronic sequencing, pyrosequencing, Maxam-Gilbert sequencing, chain termination (e.g., Sanger) sequencing, +S sequencing, or sequencing by synthesis.

[0082] Expression Systems

[0083] Provided herein are libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains, wherein the libraries have improved specificity, stability, expression, folding, or downstream activity. In some instances, libraries described herein are used for screening and analysis. [0084] Provided herein are libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains, wherein the nucleic acid libraries are used for screening and analysis. In some instances, screening and analysis comprises in vitro, in vivo, or ex vivo assays. Cells for screening include primary cells taken from living subjects or cell lines. Cells may be from prokaryotes (e.g., bacteria and fungi) or eukaryotes (e.g., animals and plants). Exemplary animal cells include, without limitation, those from a mouse, rabbit, primate, and insect. In some instances, cells for screening include a cell line including, but not limited to, Chinese Hamster Ovary (CHO) cell line, human embryonic kidney (HEK) cell line, or baby hamster kidney (BHK) cell line. In some instances, nucleic acid libraries described herein may also be delivered to a multicellular organism. Exemplary multicellular organisms include, without limitation, a plant, a mouse, rabbit, primate, and insect.

[0085] Nucleic acid libraries described herein may be screened for various pharmacological or pharmacokinetic properties. In some instances, the libraries are screened using in vitro assays, in vivo assays, or ex vivo assays. For example, in vitro pharmacological or pharmacokinetic properties that are screened include, but are not limited to, binding affinity, binding specificity, and binding avidity. Exemplary in vivo pharmacological or pharmacokinetic properties of libraries described herein that are screened include, but are not limited to, therapeutic efficacy, activity, preclinical toxicity properties, clinical efficacy properties, clinical toxicity properties, immunogenicity, potency, and clinical safety properties.

[0086] Provided herein are nucleic acid libraries, wherein the nucleic acid libraries may be expressed in a vector. Expression vectors for inserting nucleic acid libraries disclosed herein may comprise eukaryotic or prokaryotic expression vectors. Exemplary expression vectors include, without limitation, mammalian expression vectors: pSF-CMV-NEO-NH2-PPT- 3XFLAG, pSF-CMV-NEO-COOH-3XFLAG, pSF-CMV-PURO-NH2-GST-TEV, pSF-OXB20- COOH-TEV-FLAG(R)-6His, pCEP4 pDEST27, pSF-CMV-Ub-KrYFP, pSF-CMV-FMDV- daGFP, pEFla-mCherry-Nl Vector, pEFla-tdTomato Vector, pSF-CMV-FMDV-Hygro, pSF- CMV-PGK-Puro, pMCP-tag(m), and pSF-CMV-PURO-NH2-CMYC; bacterial expression vectors: pSF-OXB20-BetaGal,pSF-OXB20-Fluc, pSF-OXB20, and pSF-Tac; plant expression vectors: pRI 101-AN DNA and pCambia2301; and yeast expression vectors: pTYB21 and pKLAC2, and insect vectors: pAc5.1/V5-His A and pDEST8. In some instances, the vector is pcDNA3 or pcDNA3.1.

[0087] Described herein are nucleic acid libraries that are expressed in a vector to generate a construct comprising a scaffold comprising sequences of GPCR binding domains. In some instances, a size of the construct varies. In some instances, the construct comprises at least or about 500, 600, 700, 800, 900, 1000, 1 100, 1300, 1400, 1500, 1600, 1700, 1800, 2000, 2400, 2600, 2800, 3000, 3200, 3400, 3600, 3800, 4000, 4200,4400, 4600, 4800, 5000, 6000, 7000, 8000, 9000, 10000, or more than 10000 bases. In some instances, a the construct comprises a range of about 300 to 1,000, 300 to 2,000, 300 to 3,000, 300 to 4,000, 300 to 5,000, 300 to 6,000, 300 to 7,000, 300 to 8,000, 300 to 9,000, 300 to 10,000, 1,000 to 2,000, 1,000 to 3,000, 1,000 to 4,000, 1,000 to 5,000, 1,000 to 6,000, 1,000 to 7,000, 1,000 to 8,000, 1,000 to 9,000, 1,000 to 10,000, 2,000 to 3,000, 2,000 to 4,000, 2,000 to 5,000, 2,000 to 6,000, 2,000 to 7,000, 2,000 to 8,000, 2,000 to 9,000, 2,000 to 10,000, 3,000 to 4,000, 3,000 to 5,000, 3,000 to 6,000, 3,000 to 7,000, 3,000 to 8,000, 3,000 to 9,000, 3,000 to 10,000, 4,000 to 5,000, 4,000 to 6,000, 4,000 to 7,000, 4,000 to 8,000, 4,000 to 9,000, 4,000 to 10,000, 5,000 to 6,000, 5,000 to 7,000, 5,000 to 8,000, 5,000 to 9,000, 5,000 to 10,000, 6,000 to 7,000, 6,000 to 8,000, 6,000 to 9,000, 6,000 to 10,000, 7,000 to 8,000, 7,000 to 9,000, 7,000 to 10,000, 8,000 to 9,000, 8,000 to 10,000, or 9,000 to 10,000 bases.

[0088] Provided herein are libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains, wherein the nucleic acid libraries are expressed in a cell. In some instances, the libraries are synthesized to express a reporter gene. Exemplary reporter genes include, but are not limited to, acetohydroxyacid synthase (AHAS), alkaline phosphatase (AP), beta galactosidase (LacZ), beta glucoronidase (GUS), chloramphenicol acetyltransferase (CAT), green fluorescent protein (GFP), red fluorescent protein (RFP), yellow fluorescent protein (YFP), cyan fluorescent protein (CFP), cerulean fluorescent protein, citrine fluorescent protein, orange fluorescent protein , cherry fluorescent protein, turquoise fluorescent protein, blue fluorescent protein, horseradish peroxidase (HRP), luciferase (Luc), nopaline synthase (NOS), octopine synthase (OCS), luciferase, and derivatives thereof. Methods to determine modulation of a reporter gene are well known in the art, and include, but are not limited to, fluorometric methods (e.g. fluorescence spectroscopy, Fluorescence Activated Cell Sorting (FACS), fluorescence microscopy), and antibiotic resistance determination.

[0089] Diseases and Disorders

[0090] Provided herein are GPCR binding libraries comprising nucleic acids encoding for scaffolds comprising GPCR binding domains may have therapeutic effects. In some instances, the GPCR binding libraries result in protein when translated that is used to treat a disease or disorder. In some instances, the protein is an immunoglobulin. In some instances, the protein is a peptidomimetic. Exemplary diseases include, but are not limited to, cancer, inflammatory diseases or disorders, a metabolic disease or disorder, a cardiovascular disease or disorder, a respiratory disease or disorder, pain, a digestive disease or disorder, a reproductive disease or disorder, an endocrine disease or disorder, or a neurological disease or disorder. In some instances, the cancer is a solid cancer or a hematologic cancer. In some instances, an inhibitor of GPCR glucagon like peptide 1 receptor (GLPIR) as described herein is used for treatment of a metabolic disorder. In some instances, an inhibitor of GPCR GLPIR as described herein is used for treatment of weight gain (or for inducing weight loss), treatment of obesity, or treatment of Type II diabetes. In some instances, the subject is a mammal. In some instances, the subject is a mouse, rabbit, dog, or human. Subjects treated by methods described herein may be infants, adults, or children. Pharmaceutical compositions comprising antibodies or antibody fragments as described herein may be administered intravenously or subcutaneously. In some instances, a pharmaceutical composition comprises an antibody or antibody fragment described herein comprising a CDR-H3 comprising a sequence of any one of SEQ ID NOS: 2420 to 2436. In further instances, the pharmaceutical composition is used for treatment of a metabolic disorder.

[0091] Variant Libraries

[0092] Codon variation

[0093] Variant nucleic acid libraries described herein may comprise a plurality of nucleic acids, wherein each nucleic acid encodes for a variant codon sequence compared to a reference nucleic acid sequence. In some instances, each nucleic acid of a first nucleic acid population contains a variant at a single variant site. In some instances, the first nucleic acid population contains a plurality of variants at a single variant site such that the first nucleic acid population contains more than one variant at the same variant site. The first nucleic acid population may comprise nucleic acids collectively encoding multiple codon variants at the same variant site. The first nucleic acid population may comprise nucleic acids collectively encoding up to 19 or more codons at the same position. The first nucleic acid population may comprise nucleic acids collectively encoding up to 60 variant triplets at the same position, or the first nucleic acid population may comprise nucleic acids collectively encoding up to 61 different triplets of codons at the same position. Each variant may encode for a codon that results in a different amino acid during translation. Table 3 provides a listing of each codon possible (and the representative amino acid) for a variant site.

Table 3. List of codons and amino acids Alanine A Ala GCA GCC GCG GCT

Cysteine C Cys TGC TGT

Aspartic acid D Asp GAC GAT

Glutamic acid E Glu GAA GAG

Phenylalanine F Phe TTC TTT

Glycine G Gly GGA GGC GGG GGT

Histidine H His CAC CAT

Isoleucine I Iso ATA ATC ATT

Lysine K Lys AAA AAG

Leucine L Leu TTA TTG CTA CTC CTG CTT

Methionine M Met ATG

Asparagine N Asn AAC AAT

Proline P Pro CCA CCC CCG CCT

Glutamine Q Gin CAA CAG

Arginine R Arg AGA AGG CGA CGC CGG CGT

Serine S Ser AGC AGT TCA TCC TCG TCT

Threonine T Thr ACA ACC ACG ACT

Valine V Val GTA GTC GTG GTT

Tryptophan w Trp TGG

Tyrosine Y Tyr TAC TAT

[0094] A nucleic acid population may comprise varied nucleic acids collectively encoding up to 20 codon variations at multiple positions. In such cases, each nucleic acid in the population comprises variation for codons at more than one position in the same nucleic acid. In some instances, each nucleic acid in the population comprises variation for codons at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more codons in a single nucleic acid. In some instances, each variant long nucleic acid comprises variation for codons at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more codons in a single long nucleic acid. In some instances, the variant nucleic acid population comprises variation for codons at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more codons in a single nucleic acid. In some instances, the variant nucleic acid population comprises variation for codons in at least about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more codons in a single long nucleic acid.

[0095] Highly Parallel Nucleic Acid Synthesis

[0096] Provided herein is a platform approach utilizing miniaturization, parallelization, and vertical integration of the end-to-end process from polynucleotide synthesis to gene assembly within nanowells on silicon to create a revolutionary synthesis platform. Devices described herein provide, with the same footprint as a 96-well plate, a silicon synthesis platform is capable of increasing throughput by a factor of up to 1,000 or more compared to traditional synthesis methods, with production of up to approximately 1,000,000 or more polynucleotides, or 10,000 or more genes in a single highly-parallelized run.

[0097] With the advent of next-generation sequencing, high resolution genomic data has become an important factor for studies that delve into the biological roles of various genes in both normal biology and disease pathogenesis. At the core of this research is the central dogma of molecular biology and the concept of "residue-by -residue transfer of sequential information." Genomic information encoded in the DNA is transcribed into a message that is then translated into the protein that is the active product within a given biological pathway.

[0098] Another exciting area of study is on the discovery, development and manufacturing of therapeutic molecules focused on a highly-specific cellular target. High diversity DNA sequence libraries are at the core of development pipelines for targeted therapeutics. Gene mutants are used to express proteins in a design, build, and test protein engineering cycle that ideally culminates in an optimized gene for high expression of a protein with high affinity for its therapeutic target. As an example, consider the binding pocket of a receptor. The ability to test all sequence permutations of all residues within the binding pocket simultaneously will allow for a thorough exploration, increasing chances of success. Saturation mutagenesis, in which a researcher attempts to generate all possible mutations at a specific site within the receptor, represents one approach to this development challenge. Though costly and time and labor- intensive, it enables each variant to be introduced into each position. In contrast, combinatorial mutagenesis, where a few selected positions or short stretch of DNA may be modified extensively, generates an incomplete repertoire of variants with biased representation.

[0099] To accelerate the drug development pipeline, a library with the desired variants available at the intended frequency in the right position available for testing— in other words, a precision library, enables reduced costs as well as turnaround time for screening. Provided herein are methods for synthesizing nucleic acid synthetic variant libraries which provide for precise introduction of each intended variant at the desired frequency. To the end user, this translates to the ability to not only thoroughly sample sequence space but also be able to query these hypotheses in an efficient manner, reducing cost and screening time. Genome-wide editing can elucidate important pathways, libraries where each variant and sequence permutation can be tested for optimal functionality, and thousands of genes can be used to reconstruct entire pathways and genomes to re-engineer biological systems for drug discovery.

[00100] In a first example, a drug itself can be optimized using methods described herein. For example, to improve a specified function of an antibody, a variant polynucleotide library encoding for a portion of the antibody is designed and synthesized. A variant nucleic acid library for the antibody can then be generated by processes described herein (e.g., PCR mutagenesis followed by insertion into a vector). The antibody is then expressed in a production cell line and screened for enhanced activity. Example screens include examining modulation in binding affinity to an antigen, stability, or effector function (e.g., ADCC, complement, or apoptosis). Exemplary regions to optimize the antibody include, without limitation, the Fc region, Fab region, variable region of the Fab region, constant region of the Fab region, variable domain of the heavy chain or light chain (VH or VL), and specific complementarity-determining regions (CDRs) of VH or VL.

[00101] Nucleic acid libraries synthesized by methods described herein may be expressed in various cells associated with a disease state. Cells associated with a disease state include cell lines, tissue samples, primary cells from a subject, cultured cells expanded from a subject, or cells in a model system. Exemplary model systems include, without limitation, plant and animal models of a disease state.

[00102] To identify a variant molecule associated with prevention, reduction or treatment of a disease state, a variant nucleic acid library described herein is expressed in a cell associated with a disease state, or one in which a cell a disease state can be induced. In some instances, an agent is used to induce a disease state in cells. Exemplary tools for disease state induction include, without limitation, a Cre/Lox recombination system, LPS inflammation induction, and streptozotocin to induce hypoglycemia. The cells associated with a disease state may be cells from a model system or cultured cells, as well as cells from a subject having a particular disease condition. Exemplary disease conditions include a bacterial, fungal, viral, autoimmune, or proliferative disorder (e.g., cancer). In some instances, the variant nucleic acid library is expressed in the model system, cell line, or primary cells derived from a subject, and screened for changes in at least one cellular activity. Exemplary cellular activities include, without limitation, proliferation, cycle progression, cell death, adhesion, migration, reproduction, cell signaling, energy production, oxygen utilization, metabolic activity, and aging, response to free radical damage, or any combination thereof.

[00103] Substrates

[00104] Devices used as a surface for polynucleotide synthesis may be in the form of substrates which include, without limitation, homogenous array surfaces, patterned array surfaces, channels, beads, gels, and the like. Provided herein are substrates comprising a plurality of clusters, wherein each cluster comprises a plurality of loci that support the attachment and synthesis of polynucleotides. In some instances, substrates comprise a homogenous array surface. For example, the homogenous array surface is a homogenous plate. The term "locus" as used herein refers to a discrete region on a structure which provides support for polynucleotides encoding for a single predetermined sequence to extend from the surface. In some instances, a locus is on a two dimensional surface, e.g., a substantially planar surface. In some instances, a locus is on a three-dimensional surface, e.g. , a well, microwell, channel, or post. In some instances, a surface of a locus comprises a material that is actively functionalized to attach to at least one nucleotide for polynucleotide synthesis, or preferably, a population of identical nucleotides for synthesis of a population of polynucleotides. In some instances, polynucleotide refers to a population of polynucleotides encoding for the same nucleic acid sequence. In some cases, a surface of a substrate is inclusive of one or a plurality of surfaces of a substrate. The average error rates for polynucleotides synthesized within a library described here using the systems and methods provided are often less than 1 in 1000, less than about 1 in 2000, less than about 1 in 3000 or less often without error correction.

[00105] Provided herein are surfaces that support the parallel synthesis of a plurality of polynucleotides having different predetermined sequences at addressable locations on a common support. In some instances, a substrate provides support for the synthesis of more than 50, 100, 200, 400, 600, 800, 1000, 1200, 1400, 1600, 1800, 2,000; 5,000; 10,000; 20,000; 50,000;

100,000; 200,000; 300,000; 400,000; 500,000; 600,000; 700,000; 800,000; 900,000; 1,000,000; 1,200,000; 1,400,000; 1,600,000; 1,800,000; 2,000,000; 2,500,000; 3,000,000; 3,500,000;

4,000,000; 4,500,000; 5,000,000; 10,000,000 or more non-identical polynucleotides. In some cases, the surfaces provide support for the synthesis of more than 50, 100, 200, 400, 600, 800, 1000, 1200, 1400, 1600, 1800, 2,000; 5,000; 10,000; 20,000; 50,000; 100,000; 200,000; 300,000; 400,000; 500,000; 600,000; 700,000; 800,000; 900,000; 1,000,000; 1,200,000; 1,400,000;

1,600,000; 1,800,000; 2,000,000; 2,500,000; 3,000,000; 3,500,000; 4,000,000; 4,500,000;

5,000,000; 10,000,000 or more polynucleotides encoding for distinct sequences. In some instances, at least a portion of the polynucleotides have an identical sequence or are configured to be synthesized with an identical sequence. In some instances, the substrate provides a surface environment for the growth of polynucleotides having at least 80, 90, 100, 120, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500 bases or more.

[00106] Provided herein are methods for polynucleotide synthesis on distinct loci of a substrate, wherein each locus supports the synthesis of a population of polynucleotides. In some cases, each locus supports the synthesis of a population of polynucleotides having a different sequence than a population of polynucleotides grown on another locus. In some instances, each polynucleotide sequence is synthesized with 1, 2, 3, 4, 5, 6, 7, 8, 9 or more redundancy across different loci within the same cluster of loci on a surface for polynucleotide synthesis. In some instances, the loci of a substrate are located within a plurality of clusters. In some instances, a substrate comprises at least 10, 500, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000, 1 1000, 12000, 13000, 14000, 15000, 20000, 30000, 40000, 50000 or more clusters. In some instances, a substrate comprises more than 2,000; 5,000; 10,000; 100,000; 200,000;

300,000; 400,000; 500,000; 600,000; 700,000; 800,000; 900,000; 1,000,000; 1, 100,000;

1,200,000; 1,300,000; 1,400,000; 1,500,000; 1,600,000; 1,700,000; 1,800,000; 1,900,000;

2,000,000; 300,000; 400,000; 500,000; 600,000; 700,000; 800,000; 900,000; 1,000,000;

1,200,000; 1,400,000; 1,600,000; 1,800,000; 2,000,000; 2,500,000; 3,000,000; 3,500,000;

4,000,000; 4,500,000; 5,000,000; or 10,000,000 or more distinct loci. In some instances, a substrate comprises about 10,000 distinct loci. The amount of loci within a single cluster is varied in different instances. In some cases, each cluster includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 120, 130, 150, 200, 300, 400, 500 or more loci. In some instances, each cluster includes about 50-500 loci. In some instances, each cluster includes about 100-200 loci. In some instances, each cluster includes about 100-150 loci. In some instances, each cluster includes about 109, 121, 130 or 137 loci. In some instances, each cluster includes about 19, 20, 61, 64 or more loci. Alternatively or in combination, polynucleotide synthesis occurs on a homogenous array surface.

[00107] In some instances, the number of distinct polynucleotides synthesized on a substrate is dependent on the number of distinct loci available in the substrate. In some instances, the density of loci within a cluster or surface of a substrate is at least or about 1, 10, 25, 50, 65, 75, 100, 130, 150, 175, 200, 300, 400, 500, 1,000 or more loci per mm ² . In some cases, a substrate comprises 10-500, 25-400, 50-500, 100-500, 150-500, 10-250, 50-250, 10-200, or 50-200 mm ² . In some instances, the distance between the centers of two adjacent loci within a cluster or surface is from about 10-500, from about 10-200, or from about 10-100 um. In some instances, the distance between two centers of adjacent loci is greater than about 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 um. In some instances, the distance between the centers of two adjacent loci is less than about 200, 150, 100, 80, 70, 60, 50, 40, 30, 20 or 10 um. In some instances, each locus has a width of about 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 um. In some cases, each locus is has a width of about 0.5-100, 0.5-50, 10-75, or 0.5-50 um.

[00108] In some instances, the density of clusters within a substrate is at least or about 1 cluster per 100 mm ² , 1 cluster per 10 mm ² , 1 cluster per 5 mm ² , 1 cluster per 4 mm ² , 1 cluster per 3 mm 2 , 1 cluster per 2 mm 2 , 1 cluster per 1 mm 2 , 2 clusters per 1 mm 2 , 3 clusters per 1 mm 2 , 4 clusters per 1 mm ² , 5 clusters per 1 mm ² , 10 clusters per 1 mm ² , 50 clusters per 1 mm ² or more. In some instances, a substrate comprises from about 1 cluster per 10 mm ² to about 10 clusters per 1 mm ² . In some instances, the distance between the centers of two adjacent clusters is at least or about 50, 100, 200, 500, 1000, 2000, or 5000 um. In some cases, the distance between the centers of two adjacent clusters is between about 50-100, 50-200, 50-300, 50-500, and 100-2000 um. In some cases, the distance between the centers of two adjacent clusters is between about 0.05-50, 0.05-10, 0.05-5, 0.05-4, 0.05-3, 0.05-2, 0.1-10, 0.2-10, 0.3-10, 0.4-10, 0.5-10, 0.5-5, or 0.5-2 mm. In some cases, each cluster has a cross section of about 0.5 to about 2, about 0.5 to about 1, or about 1 to about 2 mm. In some cases, each cluster has a cross section of about 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9 or 2 mm. In some cases, each cluster has an interior cross section of about 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.15, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9 or 2 mm.

[00109] In some instances, a substrate is about the size of a standard 96 well plate, for example between about 100 and about 200 mm by between about 50 and about 150 mm. In some instances, a substrate has a diameter less than or equal to about 1000, 500, 450, 400, 300, 250, 200, 150, 100 or 50 mm. In some instances, the diameter of a substrate is between about 25-1000, 25-800, 25-600, 25-500, 25-400, 25-300, or 25-200 mm. In some instances, a substrate has a planar surface area of at least about 100; 200; 500; 1,000; 2,000; 5,000; 10,000; 12,000; 15,000; 20,000; 30,000; 40,000; 50,000 mm ² or more. In some instances, the thickness of a substrate is between about 50- 2000, 50- 1000, 100-1000, 200-1000, or 250-1000 mm.

[00110] Surface materials [00111] Substrates, devices, and reactors provided herein are fabricated from any variety of materials suitable for the methods, compositions, and systems described herein. In certain instances, substrate materials are fabricated to exhibit a low level of nucleotide binding. In some instances, substrate materials are modified to generate distinct surfaces that exhibit a high level of nucleotide binding. In some instances, substrate materials are transparent to visible and/or UV light. In some instances, substrate materials are sufficiently conductive, e.g., are able to form uniform electric fields across all or a portion of a substrate. In some instances, conductive materials are connected to an electric ground. In some instances, the substrate is heat conductive or insulated. In some instances, the materials are chemical resistant and heat resistant to support chemical or biochemical reactions, for example polynucleotide synthesis reaction processes. In some instances, a substrate comprises flexible materials. For flexible materials, materials can include, without limitation: nylon, both modified and unmodified, nitrocellulose, polypropylene, and the like. In some instances, a substrate comprises rigid materials. For rigid materials, materials can include, without limitation: glass; fuse silica; silicon, plastics (for example polytetraflouroethylene, polypropylene, polystyrene, polycarbonate, and blends thereof, and the like); metals (for example, gold, platinum, and the like). The substrate, solid support or reactors can be fabricated from a material selected from the group consisting of silicon, polystyrene, agarose, dextran, cellulosic polymers, polyacrylamides, polydimethylsiloxane (PDMS), and glass. The substrates/solid supports or the microstructures, reactors therein may be manufactured with a combination of materials listed herein or any other suitable material known in the art.

[00112] Surface Architecture

[00113] Provided herein are substrates for the methods, compositions, and systems described herein, wherein the substrates have a surface architecture suitable for the methods, compositions, and systems described herein. In some instances, a substrate comprises raised and/or lowered features. One benefit of having such features is an increase in surface area to support polynucleotide synthesis. In some instances, a substrate having raised and/or lowered features is referred to as a three-dimensional substrate. In some cases, a three-dimensional substrate comprises one or more channels. In some cases, one or more loci comprise a channel. In some cases, the channels are accessible to reagent deposition via a deposition device such as a material deposition device. In some cases, reagents and/or fluids collect in a larger well in fluid communication one or more channels. For example, a substrate comprises a plurality of channels corresponding to a plurality of loci with a cluster, and the plurality of channels are in fluid communication with one well of the cluster. In some methods, a library of polynucleotides is synthesized in a plurality of loci of a cluster. [00114] Provided herein are substrates for the methods, compositions, and systems described herein, wherein the substrates are configured for polynucleotide synthesis. In some instances, the structure is configured to allow for controlled flow and mass transfer paths for polynucleotide synthesis on a surface. In some instances, the configuration of a substrate allows for the controlled and even distribution of mass transfer paths, chemical exposure times, and/or wash efficacy during polynucleotide synthesis. In some instances, the configuration of a substrate allows for increased sweep efficiency, for example by providing sufficient volume for a growing polynucleotide such that the excluded volume by the growing polynucleotide does not take up more than 50, 45, 40, 35, 30, 25, 20, 15, 14, 13, 12, 1 1, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1%, or less of the initially available volume that is available or suitable for growing the polynucleotide. In some instances, a three-dimensional structure allows for managed flow of fluid to allow for the rapid exchange of chemical exposure.

[00115] Provided herein are substrates for the methods, compositions, and systems described herein, wherein the substrates comprise structures suitable for the methods, compositions, and systems described herein. In some instances, segregation is achieved by physical structure. In some instances, segregation is achieved by differential functionalization of the surface generating active and passive regions for polynucleotide synthesis. In some instances, differential functionalization is achieved by alternating the hydrophobicity across the substrate surface, thereby creating water contact angle effects that cause beading or wetting of the deposited reagents. Employing larger structures can decrease splashing and cross-contamination of distinct polynucleotide synthesis locations with reagents of the neighboring spots. In some cases, a device, such as a material deposition device, is used to deposit reagents to distinct polynucleotide synthesis locations. Substrates having three-dimensional features are configured in a manner that allows for the synthesis of a large number of polynucleotides (e.g., more than about 10,000) with a low error rate (e.g. , less than about 1 :500, 1 : 1000, 1 : 1500, 1 :2,000, 1 :3,000, 1 :5,000, or 1 : 10,000). In some cases, a substrate comprises features with a density of about or greater than about 1, 5, 10, 20, 30, 40, 50, 60, 70, 80, 100, 1 10, 120, 130, 140, 150, 160, 170, 180, 190, 200, 300, 400 or 500 features per mm ² .

[00116] A well of a substrate may have the same or different width, height, and/or volume as another well of the substrate. A channel of a substrate may have the same or different width, height, and/or volume as another channel of the substrate. In some instances, the diameter of a cluster or the diameter of a well comprising a cluster, or both, is between about 0.05-50, 0.05- 10, 0.05-5, 0.05-4, 0.05-3, 0.05-2, 0.05-1, 0.05-0.5, 0.05-0.1, 0.1- 10, 0.2- 10, 0.3- 10, 0.4-10, 0.5-10, 0.5-5, or 0.5-2 mm. In some instances, the diameter of a cluster or well or both is less than or about 5, 4, 3, 2, 1, 0.5, 0.1, 0.09, 0.08, 0.07, 0.06, or 0.05 mm. In some instances, the diameter of a cluster or well or both is between about 1.0 and 1.3 mm. In some instances, the diameter of a cluster or well, or both is about 1.150 mm. In some instances, the diameter of a cluster or well, or both is about 0.08 mm. The diameter of a cluster refers to clusters within a two-dimensional or three-dimensional substrate.

[00117] In some instances, the height of a well is from about 20-1000, 50-1000, 100- 1000, 200-1000, 300-1000, 400- 1000, or 500-1000 um. In some cases, the height of a well is less than about 1000, 900, 800, 700, or 600 um.

[00118] In some instances, a substrate comprises a plurality of channels corresponding to a plurality of loci within a cluster, wherein the height or depth of a channel is 5-500, 5-400, 5-300, 5-200, 5-100, 5-50, or 10-50 um. In some cases, the height of a channel is less than 100, 80, 60, 40, or 20 um.

[00119] In some instances, the diameter of a channel, locus (e.g., in a substantially planar substrate) or both channel and locus (e.g. , in a three-dimensional substrate wherein a locus corresponds to a channel) is from about 1-1000, 1-500, 1-200, 1-100, 5- 100, or 10- 100 um, for example, about 90, 80, 70, 60, 50, 40, 30, 20 or 10 um. In some instances, the diameter of a channel, locus, or both channel and locus is less than about 100, 90, 80, 70, 60, 50, 40, 30, 20 or 10 um. In some instances, the distance between the center of two adjacent channels, loci, or channels and loci is from about 1-500, 1-200, 1-100, 5-200, 5- 100, 5-50, or 5-30, for example, about 20 um.

[00120] Surface Modifications

[00121] Provided herein are methods for polynucleotide synthesis on a surface, wherein the surface comprises various surface modifications. In some instances, the surface modifications are employed for the chemical and/or physical alteration of a surface by an additive or subtractive process to change one or more chemical and/or physical properties of a substrate surface or a selected site or region of a substrate surface. For example, surface modifications include, without limitation, ( 1) changing the wetting properties of a surface, (2) functionalizing a surface, i.e., providing, modifying or substituting surface functional groups, (3) deiunctionalizing a surface, i.e., removing surface functional groups, (4) otherwise altering the chemical composition of a surface, e.g. , through etching, (5) increasing or decreasing surface roughness, (6) providing a coating on a surface, e.g. , a coating that exhibits wetting properties that are different from the wetting properties of the surface, and/or (7) depositing particulates on a surface. [00122] In some cases, the addition of a chemical layer on top of a surface (referred to as adhesion promoter) facilitates structured patterning of loci on a surface of a substrate.

Exemplary surfaces for application of adhesion promotion include, without limitation, glass, silicon, silicon dioxide and silicon nitride. In some cases, the adhesion promoter is a chemical with a high surface energy. In some instances, a second chemical layer is deposited on a surface of a substrate. In some cases, the second chemical layer has a low surface energy. In some cases, surface energy of a chemical layer coated on a surface supports localization of droplets on the surface. Depending on the patterning arrangement selected, the proximity of loci and/or area of fluid contact at the loci are alterable.

[00123] In some instances, a substrate surface, or resolved loci, onto which nucleic acids or other moieties are deposited, e.g., for polynucleotide synthesis, are smooth or substantially planar (e.g., two-dimensional) or have irregularities, such as raised or lowered features (e.g., three-dimensional features). In some instances, a substrate surface is modified with one or more different layers of compounds. Such modification layers of interest include, without limitation, inorganic and organic layers such as metals, metal oxides, polymers, small organic molecules and the like.

[00124] In some instances, resolved loci of a substrate are functionalized with one or more moieties that increase and/or decrease surface energy. In some cases, a moiety is chemically inert. In some cases, a moiety is configured to support a desired chemical reaction, for example, one or more processes in a polynucleotide synthesis reaction. The surface energy, or hydrophobicity, of a surface is a factor for determining the affinity of a nucleotide to attach onto the surface. In some instances, a method for substrate functionalization comprises: (a) providing a substrate having a surface that comprises silicon dioxide; and (b) silanizing the surface using, a suitable silanizing agent described herein or otherwise known in the art, for example, an organofunctional alkoxysilane molecule. Methods and functionalizing agents are described in U.S. Patent No. 5474796, which is herein incorporated by reference in its entirety.

[00125] In some instances, a substrate surface is functionalized by contact with a derivatizing composition that contains a mixture of silanes, under reaction conditions effective to couple the silanes to the substrate surface, typically via reactive hydrophilic moieties present on the substrate surface. Silanization generally covers a surface through self-assembly with organofunctional alkoxysilane molecules. A variety of siloxane functionalizing reagents can further be used as currently known in the art, e.g., for lowering or increasing surface energy. The organofunctional alkoxysilanes are classified according to their organic functions.

[00126] Polynucleotide Synthesis [00127] Methods of the current disclosure for polynucleotide synthesis may include processes involving phosphoramidite chemistry. In some instances, polynucleotide synthesis comprises coupling a base with phosphoramidite. Polynucleotide synthesis may comprise coupling a base by deposition of phosphoramidite under coupling conditions, wherein the same base is optionally deposited with phosphoramidite more than once, i.e., double coupling. Polynucleotide synthesis may comprise capping of unreacted sites. In some instances, capping is optional.

Polynucleotide synthesis may also comprise oxidation or an oxidation step or oxidation steps. Polynucleotide synthesis may comprise deblocking, detritylation, and sulfurization. In some instances, polynucleotide synthesis comprises either oxidation or sulfurization. In some instances, between one or each step during a polynucleotide synthesis reaction, the device is washed, for example, using tetrazole or acetonitrile. Time frames for any one step in a phosphoramidite synthesis method may be less than about 2 min, 1 min, 50 sec, 40 sec, 30 sec, 20 sec and 10 sec.

[00128] Polynucleotide synthesis using a phosphoramidite method may comprise a subsequent addition of a phosphoramidite building block (e.g., nucleoside phosphoramidite) to a growing polynucleotide chain for the formation of a phosphite triester linkage. Phosphoramidite polynucleotide synthesis proceeds in the 3' to 5 ' direction. Phosphoramidite polynucleotide synthesis allows for the controlled addition of one nucleotide to a growing nucleic acid chain per synthesis cycle. In some instances, each synthesis cycle comprises a coupling step.

Phosphoramidite coupling involves the formation of a phosphite triester linkage between an activated nucleoside phosphoramidite and a nucleoside bound to the substrate, for example, via a linker. In some instances, the nucleoside phosphoramidite is provided to the device activated. In some instances, the nucleoside phosphoramidite is provided to the device with an activator. In some instances, nucleoside phosphoramidites are provided to the device in a 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 50, 60, 70, 80, 90, 100-fold excess or more over the substrate-bound nucleosides. In some instances, the addition of nucleoside phosphoramidite is performed in an anhydrous environment, for example, in anhydrous acetonitrile. Following addition of a nucleoside phosphoramidite, the device is optionally washed. In some instances, the coupling step is repeated one or more additional times, optionally with a wash step between nucleoside phosphoramidite additions to the substrate. In some instances, a polynucleotide synthesis method used herein comprises 1, 2, 3 or more sequential coupling steps. Prior to coupling, in many cases, the nucleoside bound to the device is de-protected by removal of a protecting group, where the protecting group functions to prevent polymerization. A common protecting group is 4,4'-dimethoxytrityl (DMT). [00129] Following coupling, phosphoramidite polynucleotide synthesis methods optionally comprise a capping step. In a capping step, the growing polynucleotide is treated with a capping agent. A capping step is useful to block unreacted substrate-bound 5 ' -OH groups after coupling from further chain elongation, preventing the formation of polynucleotides with internal base deletions. Further, phosphoramidites activated with lH-tetrazole may react, to a small extent, with the 06 position of guanosine. Without being bound by theory, upon oxidation with I ₂ /water, this side product, possibly via 06-N7 migration, may undergo depurination. The apurinic sites may end up being cleaved in the course of the final deprotection of the polynucleotide thus reducing the yield of the full-length product. The 06 modifications may be removed by treatment with the capping reagent prior to oxidation with 1 ₂ /water. In some instances, inclusion of a capping step during polynucleotide synthesis decreases the error rate as compared to synthesis without capping. As an example, the capping step comprises treating the substrate- bound polynucleotide with a mixture of acetic anhydride and 1-methylimidazole. Following a capping step, the device is optionally washed.

[00130] In some instances, following addition of a nucleoside phosphoramidite, and optionally after capping and one or more wash steps, the device bound growing nucleic acid is oxidized. The oxidation step comprises the phosphite triester is oxidized into a tetracoordinated phosphate triester, a protected precursor of the naturally occurring phosphate diester

internucleoside linkage. In some instances, oxidation of the growing polynucleotide is achieved by treatment with iodine and water, optionally in the presence of a weak base (e.g., pyridine, lutidine, collidine). Oxidation may be carried out under anhydrous conditions using, e.g. tert- Butyl hydroperoxide or (l S)-(+)-( 10-camphorsulfonyl)-oxaziridine (CSO). In some methods, a capping step is performed following oxidation. A second capping step allows for device drying, as residual water from oxidation that may persist can inhibit subsequent coupling. Following oxidation, the device and growing polynucleotide is optionally washed. In some instances, the step of oxidation is substituted with a sulfurization step to obtain polynucleotide

phosphorothioates, wherein any capping steps can be performed after the sulfurization. Many reagents are capable of the efficient sulfur transfer, including but not limited to 3- (Dimethylaminomethylidene)amino)-3H-l,2,4-dithiazole-3-thion e, DDTT, 3H- l,2-benzodithiol- 3 -one 1, 1 -dioxide, also known as Beaucage reagent, and Ν,Ν,Ν'Ν'-Tetraethylthiuram disulfide (TETD).

[00131] In order for a subsequent cycle of nucleoside incorporation to occur through coupling, the protected 5 ' end of the device bound growing polynucleotide is removed so that the primary hydroxyl group is reactive with a next nucleoside phosphoramidite. In some instances, the protecting group is DMT and deblocking occurs with trichloroacetic acid in dichloromethane. Conducting detritylation for an extended time or with stronger than recommended solutions of acids may lead to increased depurination of solid support-bound polynucleotide and thus reduces the yield of the desired full-length product. Methods and compositions of the disclosure described herein provide for controlled deblocking conditions limiting undesired depurination reactions. In some instances, the device bound polynucleotide is washed after deblocking. In some instances, efficient washing after deblocking contributes to synthesized polynucleotides having a low error rate.

[00132] Methods for the synthesis of polynucleotides typically involve an iterating sequence of the following steps: application of a protected monomer to an actively functionalized surface (e.g., locus) to link with either the activated surface, a linker or with a previously deprotected monomer; deprotection of the applied monomer so that it is reactive with a subsequently applied protected monomer; and application of another protected monomer for linking. One or more intermediate steps include oxidation or sulfurization. In some instances, one or more wash steps precede or follow one or all of the steps.

[00133] Methods for phosphoramidite-based polynucleotide synthesis comprise a series of chemical steps. In some instances, one or more steps of a synthesis method involve reagent cycling, where one or more steps of the method comprise application to the device of a reagent useful for the step. For example, reagents are cycled by a series of liquid deposition and vacuum drying steps. For substrates comprising three-dimensional features such as wells, microwells, channels and the like, reagents are optionally passed through one or more regions of the device via the wells and/or channels.

[00134] Methods and systems described herein relate to polynucleotide synthesis devices for the synthesis of polynucleotides. The synthesis may be in parallel. For example, at least or about at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 1000, 10000, 50000, 75000, 100000 or more polynucleotides can be synthesized in parallel. The total number polynucleotides that may be synthesized in parallel may be from 2-100000, 3- 50000, 4- 10000, 5-1000, 6-900, 7-850, 8-800, 9-750, 10-700, 1 1-650, 12-600, 13-550, 14-500, 15-450, 16-400, 17-350, 18-300, 19-250, 20-200, 21-150,22-100, 23-50, 24-45, 25-40, 30-35. Those of skill in the art appreciate that the total number of polynucleotides synthesized in parallel may fall within any range bound by any of these values, for example 25-100. The total number of polynucleotides synthesized in parallel may fall within any range defined by any of the values serving as endpoints of the range. Total molar mass of polynucleotides synthesized within the device or the molar mass of each of the polynucleotides may be at least or at least about 10, 20, 30, 40, 50, 100, 250, 500, 750, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000, 25000, 50000, 75000, 100000 picomoles, or more. The length of each of the polynucleotides or average length of the polynucleotides within the device may be at least or about at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 100, 150, 200, 300, 400, 500 nucleotides, or more. The length of each of the polynucleotides or average length of the polynucleotides within the device may be at most or about at most 500, 400, 300, 200, 150, 100, 50, 45, 35, 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 1 1, 10 nucleotides, or less. The length of each of the

polynucleotides or average length of the polynucleotides within the device may fall from 10-500, 9-400, 1 1-300, 12-200, 13- 150, 14-100, 15-50, 16-45, 17-40, 18-35, 19-25. Those of skill in the art appreciate that the length of each of the polynucleotides or average length of the

polynucleotides within the device may fall within any range bound by any of these values, for example 100-300. The length of each of the polynucleotides or average length of the

polynucleotides within the device may fall within any range defined by any of the values serving as endpoints of the range.

[00135] Methods for polynucleotide synthesis on a surface provided herein allow for synthesis at a fast rate. As an example, at least 3, 4, 5, 6, 7, 8, 9, 10, 1 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, 50, 55, 60, 70, 80, 90, 100, 125, 150, 175, 200 nucleotides per hour, or more are synthesized. Nucleotides include adenine, guanine, thymine, cytosine, uridine building blocks, or analogs/modified versions thereof. In some instances, libraries of polynucleotides are synthesized in parallel on substrate. For example, a device comprising about or at least about 100; 1,000; 10,000; 30,000; 75,000; 100,000; 1,000,000; 2,000,000; 3,000,000; 4,000,000; or 5,000,000 resolved loci is able to support the synthesis of at least the same number of distinct polynucleotides, wherein polynucleotide encoding a distinct sequence is synthesized on a resolved locus. In some instances, a library of polynucleotides is synthesized on a device with low error rates described herein in less than about three months, two months, one month, three weeks, 15, 14, 13, 12, 1 1, 10, 9, 8, 7, 6, 5, 4, 3, 2 days, 24 hours or less. In some instances, larger nucleic acids assembled from a polynucleotide library synthesized with low error rate using the substrates and methods described herein are prepared in less than about three months, two months, one month, three weeks, 15, 14, 13, 12, 1 1, 10, 9, 8, 7, 6, 5, 4, 3, 2 days, 24 hours or less.

[00136] In some instances, methods described herein provide for generation of a library of nucleic acids comprising variant nucleic acids differing at a plurality of codon sites. In some instances, a nucleic acid may have 1 site, 2 sites, 3 sites, 4 sites, 5 sites, 6 sites, 7 sites, 8 sites, 9 sites, 10 sites, 11 sites, 12 sites, 13 sites, 14 sites, 15 sites, 16 sites, 17 sites 18 sites, 19 sites, 20 sites, 30 sites, 40 sites, 50 sites, or more of variant codon sites.

[00137] In some instances, the one or more sites of variant codon sites may be adjacent. In some instances, the one or more sites of variant codon sites may not be adjacent and separated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more codons.

[00138] In some instances, a nucleic acid may comprise multiple sites of variant codon sites, wherein all the variant codon sites are adjacent to one another, forming a stretch of variant codon sites. In some instances, a nucleic acid may comprise multiple sites of variant codon sites, wherein none the variant codon sites are adjacent to one another. In some instances, a nucleic acid may comprise multiple sites of variant codon sites, wherein some the variant codon sites are adjacent to one another, forming a stretch of variant codon sites, and some of the variant codon sites are not adjacent to one another.

[00139] Referring to the Figures, FIG. 7 illustrates an exemplary process workflow for synthesis of nucleic acids (e.g., genes) from shorter nucleic acids. The workflow is divided generally into phases: (1) de novo synthesis of a single stranded nucleic acid library, (2) joining nucleic acids to form larger fragments, (3) error correction, (4) quality control, and (5) shipment. Prior to de novo synthesis, an intended nucleic acid sequence or group of nucleic acid sequences is preselected. For example, a group of genes is preselected for generation.

[00140] Once large nucleic acids for generation are selected, a predetermined library of nucleic acids is designed for de novo synthesis. Various suitable methods are known for generating high density polynucleotide arrays. In the workflow example, a device surface layer is provided. In the example, chemistry of the surface is altered in order to improve the polynucleotide synthesis process. Areas of low surface energy are generated to repel liquid while areas of high surface energy are generated to attract liquids. The surface itself may be in the form of a planar surface or contain variations in shape, such as protrusions or micro wells which increase surface area. In the workflow example, high surface energy molecules selected serve a dual function of supporting DNA chemistry, as disclosed in International Patent Application Publication WO/2015/021080, which is herein incorporated by reference in its entirety.

[00141] In situ preparation of polynucleotide arrays is generated on a solid support and utilizes single nucleotide extension process to extend multiple oligomers in parallel. A deposition device, such as a material deposition device, is designed to release reagents in a step wise fashion such that multiple polynucleotides extend, in parallel, one residue at a time to generate oligomers with a predetermined nucleic acid sequence 702. In some instances, polynucleotides are cleaved from the surface at this stage. Cleavage includes gas cleavage, e.g., with ammonia or methylamine.

[00142] The generated polynucleotide libraries are placed in a reaction chamber. In this exemplary workflow, the reaction chamber (also referred to as "nanoreactor") is a silicon coated well, containing PCR reagents and lowered onto the polynucleotide library 703. Prior to or after the sealing 704 of the polynucleotides, a reagent is added to release the polynucleotides from the substrate. In the exemplary workflow, the polynucleotides are released subsequent to sealing of the nanoreactor 705. Once released, fragments of single stranded polynucleotides hybridize in order to span an entire long range sequence of DNA. Partial hybridization 705 is possible because each synthesized polynucleotide is designed to have a small portion overlapping with at least one other polynucleotide in the pool.

[00143] After hybridization, a PCA reaction is commenced. During the polymerase cycles, the polynucleotides anneal to complementary fragments and gaps are filled in by a polymerase. Each cycle increases the length of various fragments randomly depending on which

polynucleotides find each other. Complementarity amongst the fragments allows for forming a complete large span of double stranded DNA 706.

[00144] After PCA is complete, the nanoreactor is separated from the device 707 and positioned for interaction with a device having primers for PCR 708. After sealing, the nanoreactor is subject to PCR 709 and the larger nucleic acids are amplified. After PCR 710, the nanochamber is opened 711, error correction reagents are added 712, the chamber is sealed 713 and an error correction reaction occurs to remove mismatched base pairs and/or strands with poor complementarity from the double stranded PCR amplification products 714. The nanoreactor is opened and separated 715. Error corrected product is next subject to additional processing steps, such as PCR and molecular bar coding, and then packaged 722 for shipment 723.

[00145] In some instances, quality control measures are taken. After error correction, quality control steps include for example interaction with a wafer having sequencing primers for amplification of the error corrected product 716, sealing the wafer to a chamber containing error corrected amplification product 717, and performing an additional round of amplification 718. The nanoreactor is opened 719 and the products are pooled 720 and sequenced 721. After an acceptable quality control determination is made, the packaged product 722 is approved for shipment 723.

[00146] In some instances, a nucleic acid generate by a workflow such as that in FIG. 7 is subject to mutagenesis using overlapping primers disclosed herein. In some instances, a library of primers are generated by in situ preparation on a solid support and utilize single nucleotide extension process to extend multiple oligomers in parallel. A deposition device, such as a material deposition device, is designed to release reagents in a step wise fashion such that multiple polynucleotides extend, in parallel, one residue at a time to generate oligomers with a predetermined nucleic acid sequence 702.

[00147] Computer systems

[00148] Any of the systems described herein, may be operably linked to a computer and may be automated through a computer either locally or remotely. In various instances, the methods and systems of the disclosure may further comprise software programs on computer systems and use thereof. Accordingly, computerized control for the synchronization of the

dispense/vacuum/refill functions such as orchestrating and synchronizing the material deposition device movement, dispense action and vacuum actuation are within the bounds of the disclosure. The computer systems may be programmed to interface between the user specified base sequence and the position of a material deposition device to deliver the correct reagents to specified regions of the substrate.

[00149] The computer system 800 illustrated in FIG. 8 may be understood as a logical apparatus that can read instructions from media 811 and/or a network port 805, which can optionally be connected to server 809 having fixed media 812. The system, such as shown in FIG. 8 can include a CPU 801, disk drives 803, optional input devices such as keyboard 815 and/or mouse 816 and optional monitor 807. Data communication can be achieved through the indicated communication medium to a server at a local or a remote location. The communication medium can include any means of transmitting and/or receiving data. For example, the communication medium can be a network connection, a wireless connection or an internet connection. Such a connection can provide for communication over the World Wide Web. It is envisioned that data relating to the present disclosure can be transmitted over such networks or connections for reception and/or review by a party 822 as illustrated in FIG. 8.

[00150] FIG. 14 is a block diagram illustrating a first example architecture of a computer system 1400 that can be used in connection with example instances of the present disclosure. As depicted in FIG. 14, the example computer system can include a processor 1402 for processing instructions. Non-limiting examples of processors include: Intel XeonTM processor, AMD OpteronTM processor, Samsung 32-bit RISC ARM 1176JZ(F)-S vl .OTM processor, ARM Cortex-A8 Samsung S5PC100TM processor, ARM Cortex-A8 Apple A4TM processor, Marvell PXA 930TM processor, or a functionally-equivalent processor. Multiple threads of execution can be used for parallel processing. In some instances, multiple processors or processors with multiple cores can also be used, whether in a single computer system, in a cluster, or distributed across systems over a network comprising a plurality of computers, cell phones, and/or personal data assistant devices.

[00151] As illustrated in FIG. 9, a high speed cache 904 can be connected to, or incorporated in, the processor 902 to provide a high speed memory for instructions or data that have been recently, or are frequently, used by processor 902. The processor 902 is connected to a north bridge 906 by a processor bus 908. The north bridge 906 is connected to random access memory (RAM) 910 by a memory bus 912 and manages access to the RAM 910 by the processor 902. The north bridge 906 is also connected to a south bridge 914 by a chipset bus 916. The south bridge 914 is, in turn, connected to a peripheral bus 918. The peripheral bus can be, for example, PCI, PCI-X, PCI Express, or other peripheral bus. The north bridge and south bridge are often referred to as a processor chipset and manage data transfer between the processor, RAM, and peripheral components on the peripheral bus 918. In some alternative architectures, the functionality of the north bridge can be incorporated into the processor instead of using a separate north bridge chip. In some instances, system 900 can include an accelerator card 922 attached to the peripheral bus 918. The accelerator can include field programmable gate arrays (FPGAs) or other hardware for accelerating certain processing. For example, an accelerator can be used for adaptive data restructuring or to evaluate algebraic expressions used in extended set processing.

[00152] Software and data are stored in external storage 924 and can be loaded into RAM 910 and/or cache 904 for use by the processor. The system 900 includes an operating system for managing system resources; non-limiting examples of operating systems include: Linux, WindowsTM, MACOSTM, BlackBerry OSTM, iOSTM, and other functionally-equivalent operating systems, as well as application software running on top of the operating system for managing data storage and optimization in accordance with example instances of the present disclosure. In this example, system 900 also includes network interface cards (NICs) 920 and 921 connected to the peripheral bus for providing network interfaces to external storage, such as Network Attached Storage (NAS) and other computer systems that can be used for distributed parallel processing.

[00153] FIG. 10 is a diagram showing a network 1000 with a plurality of computer systems 1002a, and 1002b, a plurality of cell phones and personal data assistants 1002c, and Network Attached Storage (NAS) 1004a, and 1004b. In example instances, systems 1002a, 1002b, and 1002c can manage data storage and optimize data access for data stored in Network Attached Storage (NAS) 1004a and 1004b. A mathematical model can be used for the data and be evaluated using distributed parallel processing across computer systems 1002a, and 1002b, and cell phone and personal data assistant systems 1002c. Computer systems 1002a, and 1002b, and cell phone and personal data assistant systems 1002c can also provide parallel processing for adaptive data restructuring of the data stored in Network Attached Storage (NAS) 1004a and 1004b. FIG. 10 illustrates an example only, and a wide variety of other computer architectures and systems can be used in conjunction with the various instances of the present disclosure. For example, a blade server can be used to provide parallel processing. Processor blades can be connected through a back plane to provide parallel processing. Storage can also be connected to the back plane or as Network Attached Storage (NAS) through a separate network interface. In some example instances, processors can maintain separate memory spaces and transmit data through network interfaces, back plane or other connectors for parallel processing by other processors. In other instances, some or all of the processors can use a shared virtual address memory space.

[00154] FIG. 11 is a block diagram of a multiprocessor computer system 1100 using a shared virtual address memory space in accordance with an example instance. The system includes a plurality of processors 1102a-f that can access a shared memory subsystem 1104. The system incorporates a plurality of programmable hardware memory algorithm processors (MAPs)

1106a-f in the memory subsystem 1104. Each MAP 1106a-f can comprise a memory 1108a-f and one or more field programmable gate arrays (FPGAs) lllOa-f. The MAP provides a configurable functional unit and particular algorithms or portions of algorithms can be provided to the FPGAs lllOa-f for processing in close coordination with a respective processor. For example, the MAPs can be used to evaluate algebraic expressions regarding the data model and to perform adaptive data restructuring in example instances. In this example, each MAP is globally accessible by all of the processors for these purposes. In one configuration, each MAP can use Direct Memory Access (DMA) to access an associated memory 1108a-f, allowing it to execute tasks independently of, and asynchronously from the respective microprocessor 1102a-f. In this configuration, a MAP can feed results directly to another MAP for pipelining and parallel execution of algorithms.

[00155] The above computer architectures and systems are examples only, and a wide variety of other computer, cell phone, and personal data assistant architectures and systems can be used in connection with example instances, including systems using any combination of general processors, co-processors, FPGAs and other programmable logic devices, system on chips (SOCs), application specific integrated circuits (ASICs), and other processing and logic elements. In some instances, all or part of the computer system can be implemented in software or hardware. Any variety of data storage media can be used in connection with example instances, including random access memory, hard drives, flash memory, tape drives, disk arrays, Network Attached Storage (NAS) and other local or distributed data storage devices and systems.

[00156] In example instances, the computer system can be implemented using software modules executing on any of the above or other computer architectures and systems. In other instances, the functions of the system can be implemented partially or completely in firmware, programmable logic devices such as field programmable gate arrays (FPGAs) as referenced in FIG. 9, system on chips (SOCs), application specific integrated circuits (ASICs), or other processing and logic elements. For example, the Set Processor and Optimizer can be implemented with hardware acceleration through the use of a hardware accelerator card, such as accelerator card 922 illustrated in FIG. 9.

[00157] The following examples are set forth to illustrate more clearly the principle and practice of embodiments disclosed herein to those skilled in the art and are not to be construed as limiting the scope of any claimed embodiments. Unless otherwise stated, all parts and percentages are on a weight basis.

EXAMPLES

[00158] The following examples are given for the purpose of illustrating various embodiments of the disclosure and are not meant to limit the present disclosure in any fashion. The present examples, along with the methods described herein are presently representative of preferred embodiments, are exemplary, and are not intended as limitations on the scope of the disclosure. Changes therein and other uses which are encompassed within the spirit of the disclosure as defined by the scope of the claims will occur to those skilled in the art.

[00159] Example 1: Functionalization of a device surface

[00160] A device was functionalized to support the attachment and synthesis of a library of polynucleotides. The device surface was first wet cleaned using a piranha solution comprising 90% H ₂ SO ₄ and 10% H ₂ O ₂ for 20 minutes. The device was rinsed in several beakers with DI water, held under a DI water gooseneck faucet for 5 min, and dried with N ₂ . The device was subsequently soaked in NH ₄ OH (1 : 100; 3 mL:300 mL) for 5 min, rinsed with DI water using a handgun, soaked in three successive beakers with DI water for 1 min each, and then rinsed again with DI water using the handgun. The device was then plasma cleaned by exposing the device surface to O ₂ . A SAMCO PC-300 instrument was used to plasma etch O ₂ at 250 watts for 1 min in downstream mode. [00161] The cleaned device surface was actively functionalized with a solution comprising N- (3-triethoxysilylpropyl)-4-hydroxybutyramide using a YES-1224P vapor deposition oven system with the following parameters: 0.5 to 1 torr, 60 min, 70 °C, 135 °C vaporizer. The device surface was resist coated using a Brewer Science 200X spin coater. SPR™ 3612 photoresist was spin coated on the device at 2500 rpm for 40 sec. The device was pre-baked for 30 min at 90 °C on a Brewer hot plate. The device was subjected to photolithography using a Karl Suss MA6 mask aligner instrument. The device was exposed for 2.2 sec and developed for 1 min in MSF 26A. Remaining developer was rinsed with the handgun and the device soaked in water for 5 min. The device was baked for 30 min at 100 °C in the oven, followed by visual inspection for lithography defects using a Nikon L200. A descum process was used to remove residual resist using the SAMCO PC-300 instrument to O ₂ plasma etch at 250 watts for 1 min.

[00162] The device surface was passively functionalized with a 100 solution of perfluorooctyltrichlorosilane mixed with \0 μΐ _^ light mineral oil. The device was placed in a chamber, pumped for 10 min, and then the valve was closed to the pump and left to stand for 10 min. The chamber was vented to air. The device was resist stripped by performing two soaks for 5 min in 500 mL NMP at 70 °C with ultrasonication at maximum power (9 on Crest system). The device was then soaked for 5 min in 500 mL isopropanol at room temperature with ultrasonication at maximum power. The device was dipped in 300 mL of 200 proof ethanol and blown dry with N ₂ . The functionalized surface was activated to serve as a support for polynucleotide synthesis.

[00163] Example 2: Synthesis of a 50-mer sequence on an oligonucleotide synthesis device

[00164] A two dimensional oligonucleotide synthesis device was assembled into a flowcell, which was connected to a flowcell (Applied Biosystems (ABI394 DNA Synthesizer"). The two- dimensional oligonucleotide synthesis device was uniformly functionalized with N-(3- TRIETHOXYSILYLPROPYL)-4-HYDROXYBUTYRAMIDE (Gelest) was used to synthesize an exemplary polynucleotide of 50 bp ("50-mer polynucleotide") using polynucleotide synthesis methods described herein.

[00165] The sequence of the 50-mer was as described in SEQ ID NO.: 48.

5AGACAATCAACCATTTGGGGTGGACAGCCTTGACCTCTAGACTTCGGCAT##TTTT T TTTTT3' (SEQ ID NO.: 48), where # denotes Thymidine -succinyl hexamide CED

phosphoramidite (CLP-2244 from ChemGenes), which is a cleavable linker enabling the release of oligos from the surface during deprotection. [00166] The synthesis was done using standard DNA syntliesis chemistry (coupling, capping, oxidation, and deblocking) according to the protocol in Table 4 and an ABI synthesizer.

Table 4: Synthesis protocols

[00167] The phosphoramidite/activator combination was delivered similar to the delivery of bulk reagents through the flowcell. No drying steps were performed as the environment stays "wet" with reagent the entire time.

[00168] The flow restrictor was removed from the ABI 394 synthesizer to enable faster flow. Without flow restrictor, flow rates for amidites (0.1M in ACN), Activator, (0.25M

Benzoylthiotetrazole ("BTT"; 30-3070-xx from GlenResearch) in ACN), and Ox (0.02M 12 in 20% pyridine, 10% water, and 70% THF) were roughly ~100uL/sec, for acetonitrile ("ACN") and capping reagents (1 : 1 mix of CapA and CapB, wherein CapA is acetic anhydride in

THF/Pyridine and CapB is 16% 1-methylimidizole in THF), roughly ~200uL/sec, and for Deblock (3% dichloroacetic acid in toluene), roughly ~300uL/sec (compared to ~50uL/sec for all reagents with flow restrictor). The time to completely push out Oxidizer was observed, the timing for chemical flow times was adjusted accordingly and an extra ACN wash was introduced between different chemicals. After polynucleotide synthesis, the chip was deprotected in gaseous ammonia overnight at 75 psi. Five drops of water were applied to the surface to recover polynucleotides. The recovered polynucleotides were then analyzed on a Bio Analyzer small RNA chip. [00169] Example 3: Synthesis of a 100-mer sequence on an oligonucleotide synthesis device

[00170] The same process as described in Example 2 for the synthesis of the 50-mer sequence was used for the synthesis of a 100-mer polynucleotide (" 100-mer polynucleotide"; 5'

CGGGATCCTTATCGTCATCGTCGTACAGATCCCGACCCATTTGCTGTCCACCAGTCA TGCTAGCCATACCATGATGATGATGATGATGAGAACCCCGCAT##TTTTTTTTTT3', where # denotes Thymidine-succinyl hexamide CED phosphoramidite (CLP-2244 from

ChemGenes); SEQ ID NO.: 49) on two different silicon chips, the first one uniformly functionalized with N-(3-TRIETHOXYSILYLPROPYL)-4-HYDROXYBUTYRAMIDE and the second one functionalized with 5/95 mix of 11-acetoxyundecyltriethoxysilane and n- decyltriethoxysilane, and the polynucleotides extracted from the surface were analyzed on a BioAnalyzer instrument.

[00171] All ten samples from the two chips were further PCR amplified using a forward (5'ATGCGGGGTTCTCATCATC3'; SEQ ID NO.: 50) and a reverse

(5 ' CGGGATCCTTATCGTC ATCG3 ' ; SEQ ID NO.: 51) primer in a 50uL PCR mix (25uL NEB Q5 mastermix, 2.5uL lOuM Forward primer, 2.5uL lOuM Reverse primer, luL polynucleotide extracted from the surface, and water up to 50uL) using the following thermalcycling program:

98 °C, 30 sec

98 °C, 10 sec; 63 °C, 10 sec; 72 °C, 10 sec; repeat 12 cycles

72 °C, 2min

[00172] The PCR products were also run on a BioAnalyzer, demonstrating sharp peaks at the 100-mer position. Next, the PCR amplified samples were cloned, and Sanger sequenced. Table 5 summarizes the results from the Sanger sequencing for samples taken from spots 1-5 from chip 1 and for samples taken from spots 6-10 from chip 2.

Table 5: Sequencing results

Spot Error rate Cycle efficiency

8 1/1769 bp 99.94%

9 1/854 bp 99.88%

10 1/1451 bp 99.93%

[00173] Thus, the high quality and uniformity of the synthesized polynucleotides were repeated on two chips with different surface chemistries. Overall, 89% of the 100-mers that were sequenced were perfect sequences with no errors, corresponding to 233 out of 262.

[00174] Table 6 summarizes error characteristics for the sequences obtained from the polynucleotides samples from spots 1-10.

Table 6: Error characteristics

ROI MP MP MP MP MP MP MP MP MP MP Err:

Minus Err: ~1 Err: ~1 Err: ~1 Err: ~1 Err: ~1 Err: ~1 Err: ~1 Err: ~1 Err: ~1 ~1 in

Primer in 763 in 824 in 780 in 429 in 1525 in 1615 in 531 in 1769 in 854 1451

Error Rate

[00175] Example 4: Design of G protein-coupled receptor binding domains based on conformational ligand interactions

[00176] G protein-coupled receptor (GPCR) binding domains were designed using interaction surfaces between conformational ligands that interact with GPCRs. Analysis of the interaction surfaces between chemokines and cytokines and the GPCRs indicated that the N-terminal peptide prior to the first conformational cysteine represents the activation peptide, and the core helical and beta-turn-beta topologies mediate interactions with the extracellular domain (ECD) of the GPCR.

[00177] An additional 254 GPCR ligands were designed based on cross-searching Uniprot and IUPHAR databases. The ligands represented 112 human, 71 rat, 4 pig, 1 sheep, and 1 cow derived interaction classes. The ligands were then collapsed to the following 101 cross-species ligand sequence annotations: ADM, ADM2, Agouti -related protein, Angiotensinogen, Annexin Al, Apelin, Apelin receptor early, Appetite regulating hormone, Beta-defensin 4A, C-C motif chemokine, C-X-C motif chemokine, Calcitonin, Calcitonin gene-related peptide, Cathepsin G, Cathepsin G (Fragment), Cholecystokinin, Complement C3, Complement C5, Complement C5 (Fragment), Corticoliberin, Cortistatin, Cytokine SCM-1 beta, Endothelin-2, Endothelin-3, Eotaxin, Fractalkine, Galanin peptides, Galanin-like peptide, Gastric inhibitory polypeptide, Gastrin, Gastrin-releasing peptide, Glucagon, Growth-regulated alpha protein, Heme-binding protein 1, Humanin, Insulin-like 3, Insulin-like peptide INSL5, Interleukin-8, Islet amyloid polypeptide, Kininogen-1, Lymphotactin, Metastasis-suppressor KiSS-1, Neurokinin-B, Neuromedin-B, Neuromedin-S, Neuromedin-U, Neuropeptide B, Neuropeptide S, Neuropeptide W, Neurotensin/neuromedin N, Orexigenic neuropeptide QRFP, Orexin, Oxytocin-neurophysin 1, Pancreatic prohormone, Parathyroid hormone, Parathyroid hormone-related protein, Peptide YY, Pituitary adenylate cyclase-activating, Platelet basic protein, Platelet factor 4,

Prepronociceptin, Pro-FMRFamide-related neuropeptide FF, Pro-FMRFamide-related neuropeptide VF, Pro-MCH, Pro-neuropeptide Y, Pro-opiomelanocortin, Pro-thyrotropin releasing hormone, Proenkephalin-A, Proenkephalin-B, Progonadoliberin-1, Progonadoliberin-2, Prokineticin-1, Prokineticin-2, Prolactin-releasing peptide, Promotilin, Protachykinin-1, Protein Wnt-2, Protein Wnt-3a, Protein Wnt-4, Protein Wnt-5a, Protein Wnt-7b, Prothrombin, Proto- oncogene Wnt-1, Protooncogene Wnt-3, Putative uncharacterized protein, RCG55748, Retinoic acid receptor, Secretin, Somatoliberin, Somatostatin, Stromal cell-derived factor, T-kininogen 2, Tuberoiniundibular peptide of, Urocortin, Urocortin-2, Urocortin-3, Urotensin-2, Urotensin-2B, VEGF coregulated chemokine, VIP peptides, and Vasopressin-neurophysin 2-copeptin.

[00178] Structural analysis of the ligands was performed and indicated that a majority of them comprised the N-terminal activation peptide of about 11 amino acids. Motif variants were then created by trimming back the N-terminal activation peptide. As seen in Table 7, an exemplary set of variants were created based on the N-terminal activation peptide for stromal derived factor- 1. The motif variants were also placed combinatorially at multiple positions in the CDR- H3. A total of 1016 motifs were extracted for placement in the CDR-H3. In addition, the motif variants were provided with variably boundary placement and with 5-20 substring variants that were also placed in the CDR-H3.

Table 7. Variant amino acid sequences for stromal derived factor-1

[00179] As seen in Table 8, an exemplary set of variants were created for interleukin-8 based on the following sequence:

MTSKLAVALLAAFLISAALCEGAVLPRSAKELRCQCIKTYSKPFHPKFIKELRVIESGPH CANTEIIVKLSDGRELCLDPKENWVQRVVEKFLKRAENS (SEQ ID NO: 52).

Table 8. Variant amino acid sequences for inteleukin-8

[00180] Example 5: Design of G protein-coupled receptor binding domains based on peptide ligand interactions

[00181] GPCR binding domains were designed based on interaction surfaces between peptide ligands that interact with class B GPCRs. About 66 different ligands were used and include the following ligand sequence annotations: Adrenomedullin, Amylin, Angiotensin, Angiotensin I, Angiotensin II, Angiotensin III, Apelin, Apstatin, Big Endothelin, Big Gastrin, Bradykinin, Caerulein, Calcitonin, Calcitonin Gene Related Peptide, CGRP, Cholecystokinin, Endothelin, Endothelin 1, Endothelin 2, Endothelin 3, GIP, GIPs, GLP, Galanin, Gastrin, Ghrelin, Glucagon, IAPP, Kisspeptin, Mca, Metasti, Neuromedin, Neuromedin N, Neuropeptide, Neuropeptide F, Neuropeptide Y, Neurotensin, Nociceptin, Orexin, Orexin A, Orphanin, Oxytocin, Oxytocin Galanin, PACAP, PACAPs, PAR (Protease Activated Receptor) Peptides, PAR-1 Agonist, Pramlintide, Scyliorhinin I, Secretin, Senktide, Somatostatin, Somatostatin 14, Somatostatin 28, Substance P, Urotensin II, VIP, VIPs, Vasopressin, Xenin, cinnamoyl, furoyl, gastrin, holecystokinin, a-Mating Factor Pheromone. It was observed that the peptides formed a stabilized interaction with the GPCR extracellular domain (ECD).

[00182] Motif variants were generated based on the interaction surface of the peptides with the ECD as well as with the N-terminal GPCR ligand interaction surface. This was done using structural modeling. Exemplary motif variants were created based on glucagon like peptide's interaction with its GPCR as seen in Table 9. The motif variant sequences were generated using the following sequence from glucagon like peptide:

HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG (SEQ ID NO: 73).

Table 9. Variant amino acid sequences for glucagon like peptide

[00183] Example 6: Design of G protein-coupled receptor binding domains based on small molecule interactions [00184] GPCR binding domains were designed based on interaction surfaces between small molecule ligands that interact with GPCRs. By analyzing multiple GPCR ligands, an amino acid library of Tyr, Pro, Phe, His, and Gly was designed as being able to recapitulate many of the structural contacts of these ligands. An exemplary motif variant that was generated based on these observations comprises the following sequence:

sgggg(F,G,H,P,Y)(F,G,H,P,Y)(F,G,H,P,Y)(F,G,H,P,Y)(F,G,H,P,Y) (F,G,H,P,Y)(F,G,H,P,Y)(F,G ,Η,Ρ,Υ) (SEQ ID NO: 83).

[00185] Example 7: Design of G protein-coupled binding domains based on extracellular domain interactions

[00186] GPCR binding domains were designed based on interaction surfaces on extracellular domains (ECDs) and extracellular loops (ECLs) of GPCRs. About 2,257 GPCRs from human (356), mouse (369), rat (259), cow (102), pig (60), primate, fish, fly, and over 200 other organisms were analyzed, and it was observed that ECDs provide multiple complementary contacts to other loops and helices of the GPCR at a length of 15 amino acids. Further analysis of the ECLs from the about 2,257 GPCRs and all solved structures of GPCRs demonstrated that the N-terminal ECD 1 and ECL2 comprise longer extracellular sequences and provide GPCR extracellular contacts.

[00187] Motif variants were then generated based on these sequences. Exemplary variants based on the following sequence from retinoic acid induced protein 3 (GPRC5A) were generated: EYIVLTMNRTNVNVFSELSAPRRNED (SEQ ID NO: 84). See Table 10.

Table 10. Amino acid sequences

[00188] Example 8: Design of antibody scaffolds

[00189] To generate scaffolds, structural analysis, repertoire sequencing analysis of the heavy chain, and specific analysis of heterodimer high-throughput sequencing datasets were performed. Each heavy chain was associated with each light chain scaffold. Each heavy chain scaffold was assigned 5 different long CDR-H3 loop options. Each light chain scaffold was assigned 5 different L3 scaffolds. The heavy chain CDR-H3 stems were chosen from the frequently observed long H3 loop stems (10 amino acids on the N-terminus and the C-terminus) found both across individuals and across V-gene segments. The light chain scaffold L3s were chosen from heterodimers comprising long H3s. Direct heterodimers based on information from the Protein Data Bank (PDB) and deep sequencing datasets were used in which CDR HI, H2, LI, L2, L3, and CDR-H3 stems were fixed. The various scaffolds were then formatted for display on phage to assess for expression.

[00190] Structural Analysis

[00191] About 2,017 antibody structures were analyzed from which 22 structures with long CDR-H3s of at least 25 amino acids in length were observed. The heavy chains included the following: IGHV1-69, IGHV3-30, IGHV4-49, and IGHV3-21. The light chains identified included the following: IGLV3-21, IGKV3-11, IGKV2-28, IGKV1-5, IGLV1-51, IGLV1-44, and IGKV1-13. In the analysis, four heterodimer combinations were observed multiple times including: IGHV4-59/61-IGLV3-21, IGHV3-21-IGKV2-28, IGHV1-69-IGKV3-11, and IGHV1-69-IGKV1-5. An analysis of sequences and structures identified intra-CDR-H3 disulfide bonds in a few structures with packing of bulky side chains such as tyrosine in the stem providing support for long H3 stability. Secondary structures including beta-turn-beta sheets and a "hammerhead" subdomain were also observed.

[00192] Repertoire Analysis

[00193] A repertoire analysis was performed on 1,083,875 IgM+/CD27-nai ^' ve B cell receptor (BCR) sequences and 1,433,011 CD27+ sequences obtained by unbiased 5'RACE from 12 healthy controls. The 12 healthy controls comprised equal numbers of male and female and were made up of 4 Caucasian, 4 Asian, and 4 Hispanic individuals. The repertoire analysis demonstrated that less than 1% of the human repertoire comprises BCRs with CDR-H3s longer than 21 amino acids. A V-gene bias was observed in the long CDR3 subrepertoire, with IGHV1- 69, IGHV4-34, IGHV1-18, and IGHV1-8 showing preferential enrichment in BCRs with long H3 loops. A bias against long loops was observed for IGHV3-23, IGHV4-59/61, IGHV5-51, IGHV3-48, IGHV3-53/66, IGHV3-15, IGHV3-74, IGHV3-73, IGHV3-72, and IGHV2-70. The IGHV4-34 scaffold was demonstrated to be autoreactive and had a short half-life.

[00194] Viable N-terminal and C-terminal CDR-H3 scaffold variation for long loops were also designed based on the 5'RACE reference repertoire. About 81,065 CDR-H3s of amino acid length 22 amino acids or greater were observed. By comparing across V-gene scaffolds, scaffold-specific H3 stem variation was avoided as to allow the scaffold diversity to be cloned into multiple scaffold references.

[00195] Heterodimer Analysis

[00196] Heterodimer analysis was performed on scaffolds having sequences as seen in FIGS. 12A-12C. Variant sequences and lengths of the scaffolds were assayed.

[00197] Structural Analysis [00198] Structural analysis was performed using GPCR scaffolds of variant sequences and lengths were assayed. See FIG. 13.

[00199] Example 9: Generation of GPCR antibody libraries

[00200] Based on GPCR-ligand interaction surfaces and scaffold arrangements, libraries were designed and de novo synthesized. See Examples 4-8. Referring to FIG. 5, 10 variant sequences were designed for the variable domain, heavy chain 503, 237 variant sequences were designed for the heavy chain complementarity determining region 3 507, and 44 variant sequences were designed for the variable domain, light chain 513. The fragments were synthesized as three fragments as seen in FIG. 6 following similar methods as described in Examples 1-3.

[00201] Following de novo synthesis, 10 variant sequences were generated for the variable domain, heavy chain 602, 236 variant sequences were generated for the heavy chain

complementarity determining region 3 604, and 43 variant sequences were designed for a region comprising the variable domain 606, light chain and CDR-L3 and of which 9 variants for variable domain, light chain were designed. This resulted in a library with about 10 ⁵ diversity (10 x 236 x 43). This was confirmed using next generation sequencing (NGS) with 16 million reads. As seen in FIG. 14, the normalized sequencing reads for each of the 10 variants for the variable domain, heavy chain was about 1. As seen in FIG. 15, the normalized sequencing reads for each of the 43 variants for the variable domain, light chain was about 1. As seen in FIG. 16, the normalized sequencing reads for 236 variant sequences for the heavy chain complementarity determining region 3 were about 1.

[00202] The various light and heavy chains were then tested for expression and protein folding. Referring to FIGS. 17A-17D, the 10 variant sequences for variable domain, heavy chain included the following: IGHV1-18, IGHV1-69, IGHV1-8 IGHV3-21, IGHV3-23, IGHV3- 30/33rn, IGHV3-28, IGHV3-74, IGHV4-39, and IGHV4-59/61. Of the 10 variant sequences, IGHV1-18, IGHV1-69, and IGHV3-30/33rn exhibited improved characteristics such as improved thermostability. Referring to FIGS. 18A-18F, 9 variant sequences for variable domain, light chain included the following: IGKV1-39, IGKV1-9, IGKV2-28, IGKV3-11, IGKV3-15, IGKV3-20, IGKV4-1, IGLV1-51, and IGLV2-14. Of the 9 variant sequences, IGKV1-39, IGKV3-15, IGLV1-51, and IGLV2-14 exhibited improved characteristics such as improved thermostability.

[00203] Example 10: Expression of GPCR antibody libraries in HEK293 cells

[00204] Following generation of GPCR antibody libraries as in Example 13, about 47 GPCRs were selected for screening. GPCR constructs about 1.8 kb to about 4.5 kb in size were designed in a pCDNA3.1 vector. The GPCR constructs were then synthesized following similar methods as described in Examples 2-4 including hierarchal assembly. Of the 47 GPCR constructs, 46 GPCR constructs were synthesized.

[00205] The synthesized GPCR constructs were transfected in HEK293 and assayed for expression using immunofluorescence. Referring to FIGS. 19A-19C, HEK293 cells were transfected with the GPCR constructs comprising an N-terminally hemagglutinin (HA)-tagged human Yi receptor. Following 24-48 hours of transfection, cells were washed with phosphate buffered saline (PBS) and fixed with 4% paraformaldehyde. Cells were stained using fluorescent primary antibody directed towards the HA tag or secondary antibodies comprising a fluorophore and DAPI to visualize the nuclei in blue. Referring to FIGS. 19A-19C, human Yi receptor was visualized on the cell surface in non-permeabilized cells and on the cell surface and

intracellularly in permeabilized cells.

[00206] GPCR constructs were also visualized by designing GPCR constructs comprising auto-fluorescent proteins. Referring to FIGS. 20A-20C, human Yi receptor comprised EYFP fused to its C-terminus, and human Y ₅ receptor comprised ECFP fused to its C-terminus.

HEK293 cells were transfected with human Yi receptor or co-transfected with human Yi receptor and human Y ₅ receptor. Following transfection cells were washed and fixed with 4% paraformaldehyde. Cells were stained with DAPI. Localization of human Yi receptor and human Y5 receptor were visualized by fluorescence microscopy.

[00207] Example 11 : Design of immunoglobulin library

[00208] An immunoglobulin scaffold library was designed for placement of GPCR binding domains and for improving stability for a range of GPCR binding domain encoding sequences. The immunoglobulin scaffold included a VH domain attached with a VL domain with a linker. Variant nucleic acid sequences were generated for the framework elements and CDR elements of the VH domain and VL domain. The structure of the design is shown in FIG. 21A. A full domain architecture is shown in FIG. 21B. Sequences for the leader, linker, and pIII are listed in Table 11.

Table 11. Nucleotide sequences

[00209] The VL domains that were designed include IGKV1-39, IGKV3-15, IGLV1-51, and IGLV2-14. Each of four VL domains were assembled with their respective invariant four framework elements (FWl, FW2, FW3, FW4) and variable 3 CDR (LI, L2, L3) elements. For IGKV1-39, there was 490 variants designed for LI, 420 variants designed for L2, and 824 variants designed for L3 resulting in a diversity of 1.7 x 10 ⁸ (490 * 420 * 824). For IGKV3-15, there was 490 variants designed for LI, 265 variants designed for L2, and 907 variants designed for L3 resulting in a diversity of 1.2 x 10 ⁸ (490*265*907). For IGLVl-51, there was 184 variants designed for LI, 151 variants designed for L2, and 824 variants designed for L3 resulting in a diversity of 2.3 x 10 ⁷ (184* 151*824). IGLV2-14, 967 variants designed for LI, 535 variants designed for L2, and 922 variants designed for L3 resulting in a diversity of 4.8 10 ⁸ (967*535*922). Table 12 lists the amino acid sequences and nucleotide sequences for the four framework elements (FWl, FW2, FW3, FW4) for IGLVl-51. Table 13 lists the variable 3 CDR (LI, L2, L3) elements for IGLVl-51. Variant amino acid sequences and nucleotide sequences for the four framework elements (FWl, FW2, FW3, FW4) and the variable 3 CDR (LI, L2, L3) elements were also designed for IGKV1-39, IGKV3-15, and IGLV2-14.

Table 12. Sequences for IGLVl-51 framework elements

Table 13. Sequences for IGLVl-51 CDR elements

151 SGGSFNIGN 334 TCTGGAGGCAGCTTCAATATTGGGAATAATTATGT NYVS ATCC

152 SGSTSNIGE 335 TCTGGAAGCACTTCCAACATTGGGGAGAATTATGT NYVS GTCC

153 SGSSSNIGS 336 TCTGGAAGCAGCTCCAATATTGGGAGTGATTATGT DYVS ATCC

154 SGTSSNIGS 337 TCTGGAACCAGCTCCAACATTGGGAGTAATTATGT NYVS ATCC

155 SGSSSNIGT 338 TCTGGAAGCAGCTCCAACATTGGGACTAATTTTGT NFVS ATCC

156 SGSSSNFG 339 TCTGGAAGCAGCTCCAACTTTGGGAATAATTATGT NNYVS ATCC

157 SGSTSNIGN 340 TCTGGAAGCACCTCCAACATTGGGAATAATCATGT NHVS ATCC

158 SGSSSNIGN 341 TCTGGAAGCAGCTCCAACATTGGGAATGATTTTGT DFVS ATCC

159 SGSSSDIGD 342 TCTGGAAGCAGCTCCGACATTGGCGATAATTATGT NYVS GTCC

160 SGSSSNIGK 343 TCTGGAAGCAGCTCCAACATTGGGAAATATTATGT YYVS ATCC

161 SGSSSNIGG 344 TCTGGAAGCAGCTCCAACATTGGCGGTAATTATGT NYVS ATCC

162 SGSSSNTG 345 TCTGGAAGCAGCTCCAACACTGGGAATAATTATGT NNYVS ATCC

163 SGSSSNVG 346 TCTGGAAGCAGCTCCAACGTTGGGAATAATTATGT NNYVS GTCT

164 SGSSSNIAN 347 TCTGGAAGCAGCTCCAACATTGCGAATAATTTTGT NFVS ATCC

165 SGSSSNIGN 348 TCTGGAAGCAGCTCCAACATTGGGAATGATTATGT DYVS ATCC

166 SGSTSNIEN 349 TCTGGAAGCACCTCCAATATTGAGAATAATTATGT NYVS TTCC

167 SGGSSNIGN 350 TCTGGAGGCAGCTCCAATATTGGCAATAATGATGT NDVS GTCC

168 SGSTSNIGN 351 TCTGGAAGCACCTCCAACATTGGGAATCATTATGT HYVS ATCC

169 SGSSSNIGD 352 TCAGGAAGCAGCTCCAATATTGGGGATAATGATGT NDVS ATCC

170 SGYSSNIGN 353 TCTGGATACAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

171 SGSGSNIGN 354 TCTGGAAGCGGCTCCAACATTGGAAATAATTTTGT NFVS ATCC

172 SGSSSNIW 355 TCTGGAAGCAGCTCCAACATTTGGAATAATTATGT NNYVS ATCC

173 FGSSSNIGN 356 TTTGGAAGCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

174 SGSSSNIEK 357 TCTGGAAGCAGCTCCAACATTGAGAAGAATTATGT NYVS ATCC

175 SGSRSNIGN 358 TCTGGAAGTAGATCCAATATTGGAAATTATTATGT YYVS ATCC 176 SGTKSNIG 359 TCTGGAACCAAGTCAAACATTGGGAATAATTATGT N YVS ATCT

177 SGSTSNIGN 360 TCTGGAAGCACCTCCAACATTGGGAATTATTATGT YYVS ATCC

178 SGTSSNIGN 361 TCTGGAACCAGCTCCAACATTGGGAATAATTATGT NYVA GGCC

179 PGTSSNIGN 362 CCTGGAACCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

180 SGSTSNIGI 363 TCCGGAAGCACCTCCAACATTGGGATTAATTATGT NYVS ATCC

181 SGSSSNIGS 364 TCTGGAAGCAGCTCCAACATTGGGAGTAATCTGGT NLVS ATCC

182 SGSSSNIEN 365 TCTGGAAGCAGCTCCAACATTGAGAATAATCATGT NHVS ATCC

183 SGTRSNIGN 366 TCTGGAACCAGGTCCAACATCGGCAATAATTATGT NYVS TTCG

184 SGSTSNIGD 367 TCTGGAAGCACCTCCAACATTGGGGACAATTATGT NYVS TTCC

185 SGGSSNIGK 368 TCTGGAGGCAGTTCCAACATTGGGAAGAATTTTGT NFVS ATCC

186 SGSRSDIGN 369 TCTGGAAGCAGGTCCGACATTGGGAATAATTATGT NYVS ATCC

187 SGTSSNIGN 370 TCTGGAACTAGCTCCAACATTGGGAATAATGATGT NDVS ATCC

188 SGSSSNIGS 371 TCTGGAAGCAGCTCCAACATTGGGAGTAAATATGT KYVS ATCA

189 SGSSFNIGN 372 TCTGGAAGCAGCTTCAACATTGGGAATAATTATGT NYVS ATCC

190 SGSSSNIGN 373 TCTGGAAGCAGCTCCAACATTGGGAATACTTATGT TYVS ATCC

191 SGSSSNIGD 374 TCTGGAAGCAGCTCCAATATTGGGGATAATCATGT NHVS ATCC

192 SGSSSNIGN 375 TCTGGAAGCAGCTCCAACATTGGCAATAATCATGT NHVS TTCC

193 SGSTSNIGN 376 TCTGGAAGCACCTCCAACATTGGGAATAATGATGT NDVS ATCC

194 SGSRSNVG 377 TCTGGAAGCAGATCCAACGTTGGCAATAATTATGT NNYVS TTCA

195 SGGTSNIG 378 TCCGGAGGCACCTCCAACATTGGGAAGAATTATGT KNYVS GTCT

196 SGSSSNIAD 379 TCTGGAAGCAGCTCCAACATTGCCGATAATTATGT NYVS TTCC

197 SGSSSNIGA 380 TCTGGAAGCAGCTCCAACATTGGCGCCAATTATGT NYVS ATCC

198 SGSSSNIGS 381 TCTGGAAGCAGCTCCAACATTGGGAGTAATTATGT NYVA GGCC

199 SGSSSNIGN 382 TCTGGAAGCAGCTCCAACATTGGGAACAATTTTCT NFLS CTCC

200 SGRSSNIGK 383 TCTGGAAGAAGCTCCAACATTGGGAAGAATTATGT NYVS ATCC 201 SGSSPNIGA 384 TCTGGAAGCAGCCCCAACATTGGGGCTAATTATGT NYVS ATCC

202 SGSSSNIGP 385 TCCGGAAGCAGCTCCAACATTGGGCCTAATTATGT NYVS GTCC

203 SGSSSTIGN 386 TCTGGAAGCAGCTCCACCATTGGGAATAATTATAT NYIS ATCC

204 SGSSSNIGN 387 TCTGGAAGCAGCTCCAACATTGGGAATTATTTTGT YFVS ATCC

205 SGSRSNIGN 388 TCTGGAAGCCGCTCCAACATTGGTAATAATTTTGTA NFVS TCC

206 SGGSSNIGS 389 TCTGGAGGCAGCTCCAACATTGGGAGTAATTTTGT NFVS ATCC

207 SGSSSNIGY 390 TCTGGAAGCAGCTCCAACATTGGGTATAATTATGT NYVS ATCC

208 SGTSSNIEN 391 TCTGGAACCAGCTCGAACATTGAGAACAATTATGT NYVS ATCC

209 SGSSSNIGN 392 TCTGGAAGTAGCTCCAACATTGGGAATTATTATGT YYVS ATCC

210 SGSTSNIGK 393 TCTGGAAGCACCTCCAACATTGGGAAGAATTATGT NYVS ATCC

211 SGSSSNIGT 394 TCTGGAAGCAGTTCCAACATTGGGACTTATTATGTC YYVS TCT

212 SGSSSNVG 395 TCTGGAAGCAGCTCCAACGTTGGGAAAAATTATGT KNYVS ATCT

213 SGSTSNIGD 396 TCTGGAAGCACCTCCAACATTGGGGATAATTTTGT NFVS ATCC

214 SGSTSNIGT 397 TCTGGAAGCACCTCCAACATTGGAACTAATTATGT NYVS TTCC

215 SGGTSNIG 398 TCTGGAGGTACTTCCAACATTGGGAATAATTATGT NNYVS CTCC

216 SGSYSNIGN 399 TCTGGAAGCTACTCCAATATTGGGAATAATTATGT NYVS ATCC

217 SGSSSNIED 400 TCTGGAAGCAGCTCCAACATTGAAGATAATTATGT NYVS ATCC

218 SGSSSNIGK 401 TCTGGAAGCAGCTCCAACATTGGGAAACATTATGT HYVS ATCC

219 SGSGSNIGS 402 TCCGGTTCCGGCTCAAACATTGGAAGTAATTATGT NYVS CTCC

220 SGSSSNIGN 403 TCTGGAAGCAGCTCCAACATTGGAAATAATTATAT NYIS ATCA

221 SGASSNIGN 404 TCTGGAGCCAGTTCCAACATTGGGAATAATTATGT NYVS TTCC

222 SGRTSNIGN 405 TCTGGACGCACCTCCAACATCGGGAACAATTATGT NYVS ATCC

223 SGGSSNIGS 406 TCTGGAGGCAGCTCCAATATTGGGAGTAATTACGT NYVS ATCC

224 SGSGSNIGN 407 TCTGGAAGCGGCTCCAACATTGGGAATAATTATGT NYVS ATCC

225 SGSTSNIGS 408 TCTGGAAGCACCTCCAACATTGGGAGTAATTATGT NYVS ATCC 226 SGSSSSIGN 409 TCTGGAAGCAGCTCCAGCATTGGGAATAATTATGT NYVA GGCG

227 SGSSSNLG 410 TCTGGAAGCAGTTCCAACCTTGGAAATAATTATGT N YVS ATCC

228 SGTSSNIGK 411 TCTGGAACCAGCTCCAACATTGGGAAAAATTATGT NYVS ATCC

229 SGSSSDIGN 412 TCTGGAAGCAGCTCCGATATTGGGAACAAGTATAT KYIS ATCC

230 SGSSSNIGS 413 TCTGGAAGCAGCTCCAACATTGGAAGTAATTACAT NYIS ATCC

231 SGSTSNIGA 414 TCTGGAAGCACCTCCAACATTGGGGCTAACTATGT NYVS GTCC

232 SGSSSNIGN 415 TCTGGAAGCAGCTCCAACATTGGGAATAAGTATGT KYVS ATCC

233 SGSSSNIGN 416 TCTGGAAGCAGCTCCAACATTGGGAATAATTATGG NYGS ATCC

234 SGSTSNIAN 417 TCTGGAAGCACCTCCAACATTGCGAATAATTATGT NYVS ATCC

235 SGSYSNIGS 418 TCTGGAAGCTACTCCAATATTGGGAGTAATTATGT NYVS ATCC

236 SGSSSNIGS 419 TCTGGAAGCAGCTCCAACATTGGGAGTAATTTTGT NFVS ATCC

237 SGSSSNLEN 420 TCTGGAAGCAGCTCCAATCTTGAGAATAATTATGT NYVS ATCC

238 SGSISNIGS 421 TCTGGAAGCATCTCCAATATTGGCAGTAATTATGT NYVS ATCC

239 SGSSSDIGS 422 TCTGGAAGCAGCTCCGACATTGGGAGTAATTATGT NYVS ATCC

240 SGSSSNIGT 423 TCTGGAAGCAGCTCCAACATTGGGACTAATTATGT NYVS ATCC

241 SGSSSNIGK 424 TCTGGAAGCAGCTCCAACATTGGGAAGAATTTTGT NFVS ATCC

242 SGSSSNIGN 425 TCTGGAAGCAGCTCCAACATTGGGAATAATTTTAT NFIS ATCC

243 SGGSSNIGN 426 TCTGGAGGCAGCTCCAACATTGGCAATAATTATGT NYVS TTCC

244 SGSSSNIGE 427 TCTGGAAGCAGCTCCAACATTGGGGAGAATTATGT NYVS ATCC

245 SGSSSNIGN 428 TCTGGAAGCAGCTCCAATATTGGGAATAATTTTGT NFVA GGCC

246 SGGSSNIGN 429 TCTGGAGGCAGCTCCAACATTGGGAATAATTATGT NYVA AGCC

247 SGSSSHIGN 430 TCTGGAAGCAGCTCCCACATTGGAAATAATTATGT NYVS ATCC

248 SGSSSNIGS 431 TCTGGAAGCAGCTCCAATATTGGAAGTAATGATGT NDVS ATCG

249 SGSSSNIGN 432 TCTGGAAGCAGCTCCAACATTGGGAATAATTATGT NYVT AACC

250 SGSSSNIGN 433 TCTGGAAGCAGCTCCAACATTGGGAATAATCCTGT NPVS ATCC 251 SGGSSNIGN 434 TCTGGAGGCAGCTCCAATATTGGGAATCATTATGT HYVS ATCC

252 SGTSSNIGN 435 TCTGGAACCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

253 SGSSSNIGS 436 TCTGGAAGCAGCTCCAACATTGGAAGTAATTATGT NYVS CTCG

254 SGGTSNIGS 437 TCTGGAGGCACCTCCAACATTGGAAGTAATTATGT NYVS ATCC

255 SGSKSNIGN 438 TCTGGAAGCAAGTCCAACATTGGGAATAATTATGT NYVS ATCC

256 SGRSSNIGN 439 TCTGGAAGAAGCTCCAACATTGGGAATAATTATGT NYVS ATCG

257 SGSSSNVG 440 TCTGGAAGCAGCTCCAACGTTGGGAGTAATTATGT SNYVS TTCC

258 SGSTSNIGN 441 TCTGGAAGCACCTCCAATATTGGGAATAATTTTGT NFVS ATCC

259 SGSNFNIGN 442 TCTGGAAGCAACTTCAACATTGGGAATAATTATGT NYVS CTCC

260 SGSTSNIGY 443 TCTGGAAGCACCTCCAATATTGGATATAATTATGT NYVS ATCC

261 SGSSSNIVS 444 TCTGGAAGCAGCTCCAATATTGTAAGTAATTATGT NYVS ATCC

262 SGTSSNIGN 445 TCTGGAACCAGCTCCAACATTGGGAATAATTTTGT NFVS ATCC

263 SGSSSNIGR 446 TCTGGAAGCAGCTCCAACATTGGGAGGAATTTTGT NFVS GTCC

264 SGTTSNIGN 447 TCTGGAACGACCTCCAACATTGGGAATAATTATGT NYVS CTCC

265 SGSSSNIGN 448 TCTGGAAGCAGCTCCAACATTGGGAATAATGATGT NDVS ATCC

266 SGSSSNIGN 449 TCTGGAAGCAGCTCCAACATTGGGAATCATGATGT HDVS ATCC

267 SGSSSNIGS 450 TCTGGAAGCAGCTCCAACATTGGAAGTAGTCATGT SHVS ATCC

268 SGSSSNIGI 451 TCTGGAAGCAGCTCCAACATTGGGATTCATTATGT HYVS ATCC

269 SGGGSNIG 452 TCTGGAGGCGGCTCCAACATTGGCTATAATTATGT YNYVS CTCC

270 SGSSSNIGD 453 TCTGGAAGCAGCTCCAACATTGGGGATCATTATGT HYVS GTCG

271 SGSSSNLG 454 TCTGGAAGCAGCTCCAACCTTGGGAAGAATTATGT KNYVS ATCT

272 SGSSSNIGD 455 TCTGGAAGCAGCTCCAACATTGGCGATAATTTTGT NFVS ATCC

273 SGSTSNIEK 456 TCTGGAAGCACCTCCAACATTGAGAAAAACTATGT NYVS ATCG

274 SGSSSNIGK 457 TCTGGAAGCAGCTCCAACATTGGGAAGGATTATGT DYVS ATCC

275 SGSSSNIGK 458 TCTGGAAGCAGCTCCAACATTGGGAAGAATTATGT NYVS ATCC 276 SGSSSNIGN 459 TCTGGAAGCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

277 SGSSSNIGN 460 TCTGGAAGCAGCTCCAACATTGGGAATAATTATGC NYAS CTCC

278 SGISSNIGN 461 TCTGGAATCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

279 TGSSSNIGN 462 ACTGGAAGCAGCTCCAACATTGGGAATAATTATGT NYVS ATCC

280 SGTSSNIGN 463 TCTGGAACCAGCTCCAACATTGGGAATAATCATGT NHVS TTCC

281 SGSRSNIGK 464 TCTGGAAGTCGTTCCAACATTGGGAAAAATTATGT NYVS ATCC

IGLV1-51-L2

465 DNNKRPP 616 GACAATAATAAGCGACCCCCA

466 ENNRRPS 617 GAGAATAATAGGCGACCCTCA

467 DNNKQPS 618 GACAATAATAAGCAACCCTCA

468 DNNKRPL 619 GACAATAACAAGCGACCCTTG

469 DNDKRPA 620 GACAATGATAAGCGACCCGCA

470 DNHERPS 621 GACAATCATGAGCGACCCTCA

471 ENRKRPS 622 GAAAACCGTAAGCGACCCTCA

472 DNDQRPS 623 GACAATGATCAGCGACCCTCA

473 ENYKRPS 624 GAGAATTATAAGCGACCCTCA

474 ENTKRPS 625 GAAAATACTAAGCGACCCTCA

475 DTEKRPS 626 GACACTGAGAAGAGGCCCTCA

476 DNDKRPP 627 GACAATGATAAGCGACCCCCA

477 DHNKRPS 628 GACCATAATAAGCGACCCTCA

478 GNNERPS 629 GGCAATAATGAGCGACCCTCA

479 DTSKRPS 630 GACACTAGTAAGCGACCCTCA

480 EYNKRPS 631 GAATATAATAAGCGCCCCTCA

481 ENIKRPS 632 GAAAATATTAAGCGACCCTCA

482 DNVKRPS 633 GACAATGTTAAGCGACCCTCA

483 ENDKRSS 634 GAAAACGATAAACGATCCTCA

484 ENNKRHS 635 GAAAATAATAAGCGACACTCA

485 GNDQRPS 636 GGAAATGATCAGCGACCCTCA

486 DNDRRPS 637 GACAATGATAGGCGACCCTCA

487 DNHKRPS 638 GACAATCATAAGCGGCCCTCA

488 DNNDRPS 639 GACAATAATGACCGACCCTCA

489 ENNQRPS 640 GAGAATAATCAGCGACCCTCA

490 DNNQRPS 641 GACAATAATCAGCGACCCTCA

491 ENVKRPS 642 GAGAATGTTAAGCGACCCTCA

492 DTYKRPS 643 GACACTTATAAGAGACCCTCA

493 NNNNRPS 644 AACAATAATAACCGACCCTCA

494 GNNNRPS 645 GGCAATAATAATCGACCCTCA

495 ENDQRPS 646 GAAAATGATCAGCGACCCTCA

496 DNNKRAS 647 GACAATAATAAGCGAGCCTCA

497 DNDKRPL 648 GACAATGATAAGCGACCCTTA

498 DTDERPS 649 GACACTGATGAGCGACCTTCA 499 DNRKRPS 650 GACAATAGGAAGCGACCCTCA

500 DNDARPS 651 GACAATGATGCTCGACCCTCA

501 DNNKRLS 652 GACAATAATAAGCGACTCTCA

502 DNDKRAS 653 GACAATGATAAGCGAGCCTCA

503 DNTERPS 654 GACAATACTGAGCGACCCTCA

504 DNNIRPS 655 GACAATAATATTCGACCCTCA

505 DNKRRPS 656 GACAATAAGAGGCGACCCTCA

506 DDNNRPS 657 GACGATAATAACCGACCCTCA

507 ANNRRPS 658 GCGAATAATCGACGACCCTCA

508 DNDKRLS 659 GACAATGATAAGCGACTGTCA

509 DNNKRPA 660 GACAATAATAAGCGACCCGCA

510 DNYRRPS 661 GACAATTATAGACGTCCCTCA

511 ANDQRPS 662 GCCAATGATCAGCGACCCTCA

512 DNDKRRS 663 GACAATGATAAGCGACGCTCA

513 DKNERPS 664 GACAAGAATGAGCGACCCTCA

514 DNKERPS 665 GACAATAAGGAGCGACCCTCA

515 DN KGPS 666 GACAATAATAAGGGACCCTCA

516 ENDRRPS 667 GAAAATGATAGACGACCCTCA

517 ENDERPS 668 GAAAATGATGAGCGACCCTCA

518 QNNKRPS 669 CAAAATAATAAGCGACCCTCA

519 DNRERPS 670 GACAATCGTGAGCGACCCTCA

520 DNNRRPS 671 GACAATAATAGACGACCCTCA

521 GNNRRPS 672 GGAAATAATAGGCGACCCTCA

522 DNDNRPS 673 GACAATGATAACCGACCCTCA

523 EDNKRPS 674 GAAGATAATAAGCGACCCTCA

524 DDDERPS 675 GACGATGATGAGCGGCCCTCA

525 ASNKRPS 676 GCAAGTAATAAGCGACCCTCA

526 DN KRSS 677 GACAATAATAAGCGATCCTCA

527 QNNERPS 678 CAAAATAATGAGCGACCCTCA

528 DDDRRPS 679 GACGATGATAGGCGACCCTCA

529 NNDKRPS 680 AACAATGATAAGCGACCCTCA

530 DN RPS 681 GACAATAATAACCGACCCTCA

531 DN VRPS 682 GACAATAATGTGCGACCCTCA

532 EN ERPS 683 GAAAATAATGAGCGACCCTCA

533 DNNHRPS 684 GACAATAATCACCGACCCTCA

534 DNDERPS 685 GACAATGATGAGCGCCCCTCG

535 DNIRRPS 686 GACAATATCCGGCGACCCTCA

536 DFNKRPS 687 GACTTTAATAAGCGACCCTCA

537 ETNKRPS 688 GAAACTAATAAGCGACCCTCA

538 NDNKRPS 689 AACGATAATAAGCGACCCTCA

539 DDNKRPS 690 GACGATAATAAGCGACCCTCA

540 DNYKRPS 691 GACAATTATAAGCGACCCTCA

541 HNNKRPS 692 CACAATAATAAGCGACCCTCA

542 DNHQRPS 693 GACAATCATCAGCGACCCTCA

543 DNYKRAS 694 GACAATTATAAGCGAGCCTCA

544 DNIKRPS 695 GACAATATTAAGCGACCCTCA 545 DTHKRPS 696 GACACTCATAAGCGACCCTCA

546 DTNRRPS 697 GACACTAATAGGCGACCCTCT

547 DTNQRPS 698 GACACTAATCAGCGACCCTCA

548 ESDKRPS 699 GAAAGTGATAAGCGACCCTCA

549 DNDKRSS 700 GACAATGATAAGCGATCTTCG

550 GSNKRPS 701 GGCAGTAATAAGCGACCCTCA

551 DN KRVS 702 GACAATAACAAGCGAGTTTCA

552 N RRPS 703 AACAATAATAGGCGACCCTCA

553 DNFKRPS 704 GACAATTTTAAGCGACCCTCA

554 ENDKRPS 705 GAAAATGATAAACGACCCTCA

555 EN KRLS 706 GAAAATAATAAGCGACTCTCA

556 ADNKRPS 707 GCAGATAATAAGCGACCCTCA

557 EDNERPS 708 GAAGATAATGAGCGCCCCTCA

558 DTDQRPS 709 GACACTGATCAGCGACCCTCA

559 DNYQRPS 710 GACAATTATCAGCGACCCTCA

560 DENKRPS 711 GACGAGAATAAGCGACCCTCA

561 DTNKRPS 712 GACACTAATAAGCGACCCTCA

562 DDYRRPS 713 GACGATTATCGGCGACCCTCA

563 DNDKRHS 714 GACAACGATAAGCGGCACTCA

564 ENDNRPS 715 GAAAATGATAATCGACCCTCA

565 DDNERPS 716 GACGATAATGAGCGCCCCTCA

566 DNKKRPS 717 GACAATAAGAAGCGACCCTCA

567 DVDKRPS 718 GACGTTGATAAGCGACCCTCA

568 ENKKRPS 719 GAAAATAAAAAACGACCCTCT

569 VNDKRPS 720 GTCAATGATAAGCGACCCTCA

570 DNDHRPS 721 GACAATGATCACCGACCCTCA

571 DINKRPS 722 GACATTAATAAGCGACCCTCA

572 AN ERPS 723 GCCAATAATGAGCGACCCTCA

573 DNENRPS 724 GACAATGAAAACCGACCGTCA

574 GDDKRPS 725 GGCGATGATAAGCGACCCTCA

575 ANNQRPS 726 GCCAATAATCAGCGACCTTCA

576 DDDKRPS 727 GACGATGATAAGCGACCCTCA

577 YNNKRPS 728 TACAATAATAAGCGGCCCTCA

578 EDDKRPS 729 GAAGATGATAAGCGACCCTCA

579 EN RPS 730 GAAAACAATAACCGACCCTCG

580 DNNLRPS 731 GACAATAATCTGCGACCCTCA

581 ESNKRPS 732 GAGAGTAACAAGCGACCCTCA

582 DTDKRPS 733 GACACTGATAAGCGGCCCTCA

583 DDDQRPS 734 GACGATGATCAGCGACCCTCA

584 VN KRPS 735 GTGAATAATAAGAGACCCTCC

585 DDYKRPS 736 GACGATTATAAGCGACCCTCA

586 DNTKRPS 737 GACAATACTAAGCGACCCTCA

587 DDTERPS 738 GACGATACTGAGCGACCCTCA

588 GNDKRPS 739 GGCAATGATAAGCGACCCTCA

589 DNEKRPS 740 GACAATGAAAAGCGACCCTCA

590 DNDDRPS 741 GACAATGATGACCGACCCTCA 591 DDNRRPS 742 GACGATAATAGGCGTCCCTCA

592 GNNKRPS 743 GGCAATAATAAGCGACCCTCA

593 ANDKRPS 744 GCCAATGATAAGCGACCCTCA

594 DNNKRHS 745 GACAATAATAAGCGACACTCA

595 DDNQRPS 746 GACGACAATCAGCGACCCTCA

596 GNDRRPS 747 GGCAATGATAGGCGACCCTCA

597 DNHNRPS 748 GACAATCATAACCGACCCTCA

598 DNYERPS 749 GACAATTATGAGCGACCCTCA

599 EN KRSS 750 GAAAATAATAAGCGATCCTCA

600 DDHKRPS 751 GACGATCATAAGCGGCCCTCA

601 DNNKRRS 752 GACAATAATAAACGACGTTCA

602 DNDKRPS 753 GACAATGATAAGCGACCGTCA

603 DKNKRPS 754 GACAAGAATAAGCGACCCTCA

604 DNNKRPS 755 GACAATAATAAGCGACCCTCA

605 DIDKRPS 756 GACATTGATAAGCGACCCTCA

606 DDKKRPS 757 GACGATAAGAAGCGACCCTCA

607 ANNKRPS 758 GCCAATAATAAGCGACCCTCA

608 DNDKGPS 759 GACAATGATAAGGGACCCTCA

609 EDNRRPS 760 GAAGATAATAGGCGACCCTCA

610 ENN RPS 761 GAGAATAATAAGCGACCCTCA

611 N KRPS 762 AACAATAATAAGCGACCCTCA

612 DNNERPS 763 GACAATAATGAGCGACCCTCA

613 DNIQRPS 764 GACAATATTCAGCGACCCTCA

614 DNNYRPS 765 GACAATAATTACCGACCCTCA

615 DNYNRPS 766 GACAATTATAACCGACCCTCA

IGLV1-51-L3

767 CGTWDTSL 1591 TGCGGAACATGGGATACCAGCCTGAGTGCTGTGGT SAVVF GTTC

768 CGTWDTSL 1592 TGCGGAACATGGGATACCAGCCTGAGTGCTGGGGT SAGVF GTTC

769 CGTWDTSL 1593 TGCGGAACATGGGATACCAGCCTGAGTGCTTGGGT SAWVF GTTC

770 CGTWDRSL 1594 TGCGGAACATGGGATAGGAGCCTGAGTGCGGGGG SAGVF TGTTC

771 CGTWDRSL 1595 TGCGGAACATGGGATAGGAGCCTGAGTGCTTGGGT SAWVF ATTT

772 CGTWDTSL 1596 TGCGGAACATGGGATACCAGCCTGAGTGGTGGGGT SGGVF GTTC

773 CGTWDTSL 1597 TGCGGAACATGGGATACTAGCCTGCGTGCTGGCGT RAGVF CTTC

774 CGTWDRSL 1598 TGCGGAACATGGGATAGGAGCCTGAGTGTTTGGGT SVWVF GTTC

775 CGTWDTSL 1599 TGCGGAACATGGGATACCAGTCTGAGTGTTGTGGT SWVF CTTC

776 CGTWDTSL 1600 TGCGGAACGTGGGATACCAGCCTGAGTGCTGCGGT SAAVF GTTC

777 CGAWDTSL 1601 TGCGGAGCATGGGATACCAGCCTGAGTGCTGGAGT SAGVF GTTC 778 CATWDTSL 1602 TGCGCAACATGGGATACCAGCCTGAGTGCTGTGGT SAVVF ATTC

779 CATWDTSL 1603 TGCGCAACATGGGATACCAGCCTGAGTGCTGGTGT SAGVF GTTC

780 CGTWESSL 1604 TGTGGAACATGGGAGAGCAGCCTGAGTGCTTGGGT SAWVF GTTC

781 CGTWDTTL 1605 TGCGGAACATGGGATACCACCCTGAGTGCGGGTGT SAGVF CTTC

782 CGTWDTSL 1606 TGCGGAACATGGGATACTAGCCTGAGTGTGTGGGT SVWVF GTTC

783 CGTWDTSL 1607 TGCGGAACATGGGATACTAGCCTGAGTGTTGGGGT SVGVF GTTC

784 CGTWDTSL 1608 TGCGGAACATGGGACACCAGTCTGAGCACTGGCGT STGVF CTTC

785 CGTWDTSL 1609 TGCGGAACATGGGATACCAGCCTGAGTGGTGTGGT SGVVF CTTC

786 CGTWDTSL 1610 TGCGGAACATGGGATACCAGCCTGAGTGCTTATGT SAYVF CTTC

787 CGTWDTSL 1611 TGCGGAACATGGGATACCAGCCTGAGTGCTGAGGT SAEVF GTTC

788 CGTWDTGL 1612 TGCGGAACATGGGATACCGGCCTGAGTGCTGGGGT SAGVF ATTC

789 CGTWDRSL 1613 TGCGGAACGTGGGATAGGAGCCTGAGTGCTTATGT SAYVF CTTC

790 CGTWDRSL 1614 TGCGGAACATGGGATAGGAGCCTCAGTGCCGTGGT SAVVF ATTC

791 CGTWDNTL 1615 TGCGGAACATGGGATAACACCCTGAGTGCGTGGGT SAWVF GTTC

792 CGTWDNR 1616 TGCGGAACATGGGATAACAGGCTGAGTGCTGGGGT LSAGVF GTTC

793 CGTWDISL 1617 TGCGGAACATGGGACATCAGCCTGAGTGCTTGGGT SAWVF GTTC

794 CGTWHSSL 1618 TGCGGAACATGGCATAGCAGCCTGAGTGCTGGGGT SAGVF ATTC

795 CGTWGSSL 1619 TGCGGAACATGGGGTAGCAGTTTGAGTGCTTGGGT SAWVF GTTC

796 CGTWESSL 1620 TGCGGAACATGGGAGAGCAGCCTGAGTGGTTGGGT SGWVF GTTC

797 CGTWESSL 1621 TGCGGAACATGGGAGAGCAGCCTGAGTGCTGTGGT SAVVF TTTC

798 CGTWDYSL 1622 TGCGGAACATGGGATTACAGCCTGAGTGCTGTGGT SAVVF ATTC

799 CGTWDYSL 1623 TGCGGAACATGGGATTACAGCCTGAGTGCTGGGGT SAGVF ATTC

800 CGTWDVSL 1624 TGCGGAACATGGGATGTCAGCCTGAGTGTTGGAGT SVGVF GTTC

801 CGTWDTTL 1625 TGCGGAACATGGGATACCACCCTGAGTGCTGTGGT SAVVF TTTC

802 CGTWDTTL 1626 TGCGGAACATGGGATACCACTCTGAATATTGGGGT NIGVF GTTC 803 CGTWDTSL 1627 TGCGGAACATGGGATACCAGCCTGACTGCTGTGGT TAVVF ATTC

804 CGTWDTSL 1628 TGCGGAACCTGGGATACCAGCCTGACTGCTGCTGT TAAVF GTTC

805 CGTWDTSL 1629 TGCGGCACATGGGATACCAGCCTGAGTGTGGGGCT SVGLF ATTC

806 CGTWDTSL 1630 TGCGGAACCTGGGATACCAGCCTGAGTGGTAGGGT SGRVF GTTC

807 CGTWDTSL 1631 TGCGGAACATGGGATACCAGCCTGAGTGGTGCAGT SGAVF GTTC

808 CGTWDTSL 1632 TGCGGAACATGGGATACCAGCCTGAGTGCTGGCCT SAGLF GTTC

809 CGTWDTSL 1633 TGCGGAACATGGGATACCAGCCTGAGTGCTGGAGG SAGGVF GGTCTTC

810 CGTWDTSL 1634 TGCGGAACATGGGATACCAGCCTGCGTGCTTATGT RAYVF CTTC

811 CGTWDTSL 1635 TGCGGAACATGGGATACTAGTTTGCGTGCTTGGGT RAWVF ATTC

812 CGTWDTSL 1636 TGCGGAACATGGGATACCAGCCTGAATACTGGGGT NTGVF ATTC

813 CGTWDTSL 1637 TGCGGAACATGGGATACCAGCCTGAATATTTGGGT NIWVF GTTC

814 CGTWDTSL 1638 TGCGGAACATGGGATACAAGCCTGAATATTGGGGT NIGVF GTTC

815 CGTWDTSL 1639 TGCGGAACATGGGATACCAGCCTGATTGCTGTGGT IAVVF GTTC

816 CGTWDRSL 1640 TGCGGAACGTGGGATAGGAGCCTGAGTGGTTGGGT SGWVF GTTC

817 CGTWDNR 1641 TGCGGAACATGGGATAACAGGCTGAGTGGTTGGGT LSGWVF GTTC

818 CGTWDKSL 1642 TGCGGAACGTGGGATAAGAGCCTGAGTGCTGTGGT SAVVF CTTC

819 CGTWDKG 1643 TGCGGAACATGGGATAAAGGCCTGAGTGCTTGGGT LSAWVF GTTC

820 CGTWDISL 1644 TGCGGAACATGGGATATCAGCCTGAGTGCTGGGGT SAGVF GTTC

821 CGTWDESL 1645 TGCGGAACATGGGATGAGAGCCTGAGTGGTGGCG SGGEVVF AGGTGGTCTTC

822 CGTWDASL 1646 TGCGGAACATGGGATGCCAGCCTGAGTGCCTGGGT SAWVF GTTC

823 CGTWDAG 1647 TGCGGAACTTGGGATGCCGGCCTGAGTGCTTGGGT LSAWVF GTTC

824 CGAWDTSL 1648 TGCGGAGCATGGGATACCAGCCTGAGTGCTTGGGT SAWVF GTTC

825 CGAWDTSL 1649 TGCGGAGCATGGGATACCAGCCTGAGTGCTGTGGT SAVVF GTTC

826 CGAWDTSL 1650 TGCGGAGCATGGGATACCAGCCTGCGTGCTGGGGT RAGVF TTTC

827 CATWDTSV 1651 TGCGCAACATGGGATACCAGCGTGAGTGCTTGGGT SAWVF GTTC 828 CATWDTSL 1652 TGCGCAACATGGGATACCAGCCTGAGTGCGTGGGT SAWVF GTTC

829 CATWDNTL 1653 TGCGCAACATGGGACAACACCCTGAGTGCTGGGGT SAGVF GTTC

830 CAAWDRSL 1654 TGCGCAGCATGGGATAGGAGCCTGAGTGTTTGGGT SVWVF GTTC

831 CYTWHSSL 1655 TGCTACACATGGCATTCCAGTCTGCGTGGTGGGGT RGGVF GTTC

832 CVTWTSSP 1656 TGCGTAACGTGGACTAGTAGCCCGAGTGCTTGGGT SAWVF GTTC

833 CVTWRGG 1657 TGCGTGACATGGCGTGGTGGCCTTGTGTTGTTC LVLF

834 CVTWDTSL 1658 TGCGTAACATGGGATACCAGCCTGACTTCTGTGGT TSWL ACTC

835 CVTWDTSL 1659 TGCGTAACATGGGATACCAGCCTGAGTGTTTATTG SVYWVF GGTGTTC

836 CVTWDTSL 1660 TGCGTTACATGGGATACCAGCCTGAGTGCCTGGGT SAWVF GTTC

837 CVTWDTDL 1661 TGCGTCACATGGGATACCGACCTCAGCGTTGCGCT SVALF CTTC

838 CVTWDRSL 1662 TGCGTAACATGGGATAGGAGCCTGAGTGGTTGGGT SGWVF GTTC

839 CVTWDRSL 1663 TGCGTAACATGGGATCGCAGCCTGAGAGAGGTGTT REVLF ATTC

840 CVTWDRSL 1664 TGCGTAACATGGGATCGCAGCCTGAGAGCGGTGGT RAVVF ATTC

841 CVTWDRSL 1665 TGCGTAACATGGGACAGGAGCCTCGATGCTGGGGT DAGVF TTTC

842 CVTWDNTL 1666 TGCGTGACATGGGATAACACCCTGAGTGCTGGGGT SAGVF CTTC

843 CVTWDN 1667 TGCGTAACATGGGATAACAACCTGTTTGGTGTGGT LFGVVF CTTC

844 CVSWDTSL 1668 TGCGTATCATGGGATACCAGCCTGAGTGGTGCGGT SGAVF ATTC

845 CVSWDTSL 1669 TGCGTCTCATGGGATACCAGCCTGAGTGCTGGGGT SAGVF ATTC

846 CTTWFRTP 1670 TGCACAACATGGTTTAGGACTCCGAGTGATGTGGT SDVVF CTTC

847 CTTWFRTA 1671 TGCACAACATGGTTTAGGACTGCGAGTGATGTGGT SDVVF CTTC

848 CTTWDYGL 1672 TGCACAACGTGGGATTACGGTCTGAGTGTCGTCTT SWF C

849 CTARDTSL 1673 TGCACAGCAAGGGATACCAGCCTGAGTCCTGGCGG SPGGVF GGTCTTC

850 CSTWNTRP 1674 TGCTCAACATGGAATACGAGGCCGAGTGATGTGGT SDVVF GTTC

851 CSTWESSL 1675 TGTTCAACATGGGAGAGCAGTTTGACTACTGTGGT TTVVF CTTC

852 CSTWDTSL 1676 TGCTCAACATGGGATACCAGCCTCACTAATGTGCT TNVLF ATTC 853 CSTWDTSL 1677 TGCTCAACATGGGATACCAGCCTGAGTGGAGTAGT SGVVF CTTC

854 CSTWDHSL 1678 TGCTCAACATGGGATCACAGCCTGAAAGCTGCACT KAALF GTTC

855 CSTWDARL 1679 TGCTCAACCTGGGATGCGAGGCTGAGTGTCCGGGT SVRVF GTTC

856 CSSYTSSST 1680 TGCTCCTCATATACAAGCAGCAGCACTTGGGTGTT WVF C

857 CSSYATRG 1681 TGCAGCTCATACGCAACCCGCGGCCTTCGTGTGTT LRVLF GTTC

858 CSSWDATL 1682 TGTTCATCATGGGACGCCACCCTGAGTGTTCGCAT SVRIF ATTC

859 CQVWEGSS 1683 TGTCAGGTGTGGGAGGGTAGTAGTGATCATTGGGT DHWVF GTTC

860 CQTWDNR 1684 TGCCAAACCTGGGATAACAGACTGAGTGCTGTGGT LSAVVF GTTC

861 CQTWDHSL 1685 TGTCAAACGTGGGATCACAGCCTGCATGTTGGGGT HVGVF GTTC

862 CQSYDDIL 1686 TGCCAGTCCTATGACGACATCTTGAATGTTTGGGTC NVWVL CTT

863 CNTWDKSL 1687 TGCAATACATGGGATAAGAGTTTGACTTCTGAACT TSELF CTTC

864 CLTWDRSL 1688 TGCTTAACATGGGATCGCAGCCTGAATGTGAGGGT NVRVF GTTC

865 CLTWDHSL 1689 TGCCTAACATGGGACCACAGCCTGACTGCTTATGT TAYVF CTTC

866 CLTRDTSLS 1690 TGCTTAACAAGGGATACCAGTCTGAGTGCCCCTGT APVF GTTC

867 CKTWESGL 1691 TGCAAAACATGGGAAAGTGGCCTTAATTTTGGCCA NFGHVF CGTCTTC

868 CKTWDTSL 1692 TGCAAAACATGGGATACCAGCCTGAGTGCTGTGGT SAVVF CTTC

869 CGVWDVS 1693 TGCGGAGTCTGGGATGTCAGTCTGGGTGCTGGGGT LGAGVF GTTC

870 CGVWDTTP 1694 TGCGGAGTCTGGGATACCACCCCGAGTGCCGTTCT SAVLF TTTC

871 CGVWDTTL 1695 TGCGGAGTCTGGGATACCACCCTGAGTGCCGTTCT SAVLF TTTC

872 CGVWDTSL 1696 TGCGGAGTATGGGATACCAGCCTGGGGGTCTTC GVF

873 CGVWDTN 1697 TGCGGGGTATGGGATACCAACCTGGGTAAATGGGT LGKWVF TTTC

874 CGVWDTG 1698 TGTGGAGTTTGGGATACTGGCCTGGATGCTGGTTG LDAGWVF GGTGTTC

875 CGVWDNV 1699 TGCGGAGTGTGGGATAACGTCCTGGAGGCCTATGT LEAYVF CTTC

876 CGVWDISL 1700 TGCGGAGTCTGGGATATCAGCCTGAGTGCTAATTG SANWVF GGTGTTC

877 CGVWDHS 1701 TGCGGAGTATGGGATCACAGCCTGGGGATTTGGGC LGIWAF CTTC 878 CGVWDDIL 1702 TGCGGAGTTTGGGATGATATTCTGACTGCTGAAGT TAEVF GTTC

879 CGVRDTSL 1703 TGCGGAGTTCGGGATACCAGCCTGGGGGTCTTC GVF

880 CGTYDTSL 1704 TGCGGAACATACGATACGAGCCTGCCTGCTTGGGT PAWVF GTTT

881 CGTYDNLV 1705 TGCGGAACTTACGATAATCTTGTATTTGGTTATGTC FGYVF TTC

882 CGTYDDRL 1706 TGCGGAACATACGATGATAGACTCAGAGAGGTGTT REVF C

883 CGTWVTSL 1707 TGCGGAACGTGGGTTACCAGCCTGAGTGCTGGGGT SAGVF GTTC

884 CGTWVSSL 1708 TGCGGAACATGGGTTAGCAGCCTGACTACTGTAGT TTVVF ATTC

885 CGTWVSSL 1709 TGCGGAACATGGGTTAGCAGCCTGAACGTCTGGGT NVWVF GTTC

886 CGTWVGRF 1710 TGCGGAACATGGGTTGGCAGGTTTTGGGTATTC WVF

887 CGTWSGGP 1711 TGCGGAACATGGTCTGGCGGCCCGAGTGGCCATTG SGHWLF GTTGTTC

888 CGTWSGGL 1712 TGCGGAACATGGTCTGGCGGCCTGAGTGGCCATTG SGHWLF GTTGTTC

889 CGTWQTG 1713 TGCGGAACGTGGCAGACCGGCCGGGAGGCTGTCCT REAVLF ATTT

890 CGTWQSRL 1714 TGCGGAACGTGGCAGAGCAGGCTGAGGTGGGTGTT RWVF C

891 CGTWQSRL 1715 TGCGGAACGTGGCAGAGCAGGCTGGGGTGGGTGTT GWVF C

892 CGTWPRSL 1716 TGCGGAACATGGCCTAGGAGCCTGAGTGCTGTTTG SAVWVF GGTGTTC

893 CGTWN Y 1717 TGCGGAACATGGAATAACTACCTGAGTGCTGGCGA LSAGDVVF TGTGGTTTTC

894 CGTWLGSQ 1718 TGCGGAACATGGCTTGGCAGCCAGAGTCCTTATTG SPYWVF GGTCTTC

895 CGTWHTGL 1719 TGCGGAACATGGCATACCGGCCTGAGTGCTTATGT SAYVF CTTC

896 CGTWHSTL 1720 TGCGGAACATGGCATAGTACCCTGAGTGCTGGCCA SAGHWVF TTGGGTGTTC

897 CGTWHSSL 1721 TGCGGAACATGGCATAGTAGCCTGAGTACTTGGGT STWVF GTTC

898 CGTWHSSL 1722 TGCGGAACATGGCATAGCAGCCTGAGTGCCTATGT SAYVF CTTC

899 CGTWHSSL 1723 TGCGGAACATGGCATAGCAGCCTGAGTGCTGTGGT SAVVF ATTC

900 CGTWHSGL 1724 TGCGGAACGTGGCATTCCGGCCTGAGTGGGTGGGT SGWVF TTTC

901 CGTWHNTL 1725 TGCGGAACATGGCATAACACCCTGCGTAATGTGAT RNVIF ATTC

902 CGTWHASL 1726 TGCGGAACATGGCATGCCAGCCTGACTGCTGTGTT TAVF C 903 CGTWGWY 1727 TGCGGGACATGGGGATGGTATGGCAGCCAGAGAG GSQRGVVF GCGTCGTCTTC

904 CGTWGWY 1728 TGCGGGACATGGGGATGGTATGGCGGCCAGAGAG GGQRGVVF GCGTCGTCTTC

905 CGTWGTSL 1729 TGCGGAACCTGGGGAACCAGCCTGAGTGCTTGGGT SAWVF GTTC

906 CGTWGSSL 1730 TGCGGAACCTGGGGTAGCAGCCTGACTACTGGCCT TTGLF GTTC

907 CGTWGSSL 1731 TGCGGAACATGGGGTAGCAGCCTGACTGCCTATGT TAYVF CTTC

908 CGTWGSSL 1732 TGCGGAACATGGGGTAGCAGCCTGAGTGTTGTGTT SWF C

909 CGTWGSSL 1733 TGCGGAACATGGGGTAGCAGCCTGAGTGGTGGGGT SGGVF GTTC

910 CGTWGSSL 1734 TGCGGAACATGGGGTAGCAGCCTGAGTGCTTATTG SAYWVF GGTGTTC

911 CGTWGSSL 1735 TGCGGAACATGGGGTAGCAGCCTGAGTGCTTATGT SAYVVF GGTGTTC

912 CGTWGSSL 1736 TGCGGAACATGGGGTAGCAGCCTGAGTGCTTATGT SAYVF CTTC

913 CGTWGSSL 1737 TGCGGAACGTGGGGTAGTAGCCTGAGTGCTGTGGT SAVVF GTTC

914 CGTWGSSL 1738 TGCGGAACATGGGGTAGCAGCCTGAGTGCTCCTTA SAPYVF TGTCTTC

915 CGTWGSSL 1739 TGCGGAACATGGGGTAGCAGCCTGAGTGCCCCGGT SAPVF GTTC

916 CGTWGSSL 1740 TGCGGAACATGGGGTAGCAGCCTGAGTGCTGGGGT SAGVF GTTC

917 CGTWGSSL 1741 TGCGGAACTTGGGGTAGCAGCCTGAGTGCTGGACT SAGLF GTTC

918 CGTWGSSL 1742 TGCGGAACATGGGGTAGCAGCCTGAGTGCTGGGGC SAGALF ACTCTTC

919 CGTWGSSL 1743 TGCGGAACATGGGGCAGTAGCCTGCGTGCTTGGGT RAWVF GTTC

920 CGTWFTSL 1744 TGCGGAACCTGGTTTACTAGTCTGGCTAGTGGGGT ASGVF TTTC

921 CGTWETSL 1745 TGCGGAACTTGGGAGACCAGTCTGAGTGTCGTGGT SWVI CATC

922 CGTWETSL 1746 TGCGGAACATGGGAGACCAGCCTGAGTGGTGTCTT SGVF C

923 CGTWETSL 1747 TGCGGAACATGGGAAACCAGCCTGAGTGATTGGGT SDWVF ATTC

924 CGTWETSL 1748 TGCGGAACATGGGAGACCAGCCTGAGTGCTGGGGT SAGVF ATTC

925 CGTWETSL 1749 TGCGGAACATGGGAAACCAGCCTTAATTATGTGGC NYVAF CTTC

926 CGTWETSL 1750 TGCGGAACATGGGAGACCAGCCTGAATACTTGGTT NTWLL GCTC

927 CGTWETSE 1751 TGCGGAACATGGGAGACCAGCGAGAGTGGTAATT SGNYIF ACATCTTC 928 CGTWETRL 1752 TGCGGAACATGGGAAACCAGACTGGGTACTTGGGT GTWVI GATC

929 CGTWETQL 1753 TGCGGAACATGGGAGACCCAGTTATATTGGGTGTT YWVF C

930 CGTWETGL 1754 TGCGGAACATGGGAGACTGGCCTAAGTGCTGGAGA SAGEVF GGTGTTC

931 CGTWESTL 1755 TGCGGAACTTGGGAAAGCACCCTGAGTGTTTTCCT SVFLF ATTC

932 CGTWESSL 1756 TGCGGGACATGGGAAAGTAGCCTGACTGTTGTGGT TVVVF CTTC

933 CGTWESSL 1757 TGCGGAACATGGGAAAGTAGCCTGACTGGAGTGGT TGVVF ATTC

934 CGTWESSL 1758 TGCGGAACATGGGAAAGCAGCCTGACTGGTTTTGT TGFVF CTTC

935 CGTWESSL 1759 TGTGGAACATGGGAGAGCAGCCTGAGTGTTGGGGT SVGVF GTTC

936 CGTWESSL 1760 TGCGGAACCTGGGAAAGTAGCCTCAGTGAATGGGT SEWVF GTTC

937 CGTWESSL 1761 TGCGGAACATGGGAGAGCAGCCTGAGTGCTGTATT SAVF C

938 CGTWESSL 1762 TGCGGAACATGGGAGAGCAGCCTGAGTGCTGGTTA SAGYIF TATCTTC

939 CGTWESSL 1763 TGCGGAACATGGGAGAGCAGCCTGAGTGCTGGAGT SAGVF GTTC

940 CGTWESSL 1764 TGCGGAACATGGGAAAGCAGCCTGAGCGCTGGCCC SAGPVF GGTGTTC

941 CGTWESSL 1765 TGCGGAACATGGGAAAGCAGCCTGAGTGCTGGAG SAGGQVF GCCAGGTGTTC

942 CGTWESSL 1766 TGCGGAACATGGGAGAGCAGCCTGAGTGCCTTCGG SAFGGYVF CGGTTATGTCTTC

943 CGTWESSL 1767 TGCGGAACATGGGAAAGCAGCCTGAGGGTTTGGGT RVWVF GTTC

944 CGTWESSL 1768 TGCGGAACATGGGAAAGCAGCCTCTTTACTGGGCC FTGPWVF TTGGGTGTTC

945 CGTWESLS 1769 TGCGGAACATGGGAGAGCCTGAGTGCCACCTATGT ATYVF CTTC

946 CGTWESGL 1770 TGCGGAACATGGGAGAGCGGCCTGAGTGCTGGTGT SAGVF CTTC

947 CGTWESDF 1771 TGCGGAACATGGGAAAGCGACTTTTGGGTGTTT WVF

948 CGTWENRL 1772 TGCGGTACATGGGAAAACAGACTGAGTGCTGTGGT SAVVF CTTC

949 CGTWENRL 1773 TGCGGAACATGGGAAAACAGACTGAGTGCCGGGG SAGVF TATTC

950 CGTWEISL 1774 TGCGGAACATGGGAAATCAGCCTGACTACTTCTGT TTSVVF GGTATTC

951 CGTWEISLS 1775 TGCGGAACATGGGAAATCAGCCTGAGTACTTCTGT TSWF GGTATTC

952 CGTWEGSL 1776 TGCGGAACATGGGAAGGCAGCCTCAGTGTTGTTTT SWF C 953 CGTWEGSL 1777 TGCGGAACATGGGAAGGCAGCCTGAGGGTGTTC RVF

954 CGTWEGSL 1778 TGCGGAACATGGGAGGGCAGCCTGAGGCACGTGTT RHVF C

955 CGTWDYSP 1779 TGCGGAACATGGGATTACAGCCCTGTACGTGCTGG VRAGVF GGTGTTC

956 CGTWDYSL 1780 TGCGGAACGTGGGATTACAGCCTGAGTGTTTATCT SVYLF CTTC

957 CGTWDYSL 1781 TGCGGAACATGGGATTACAGCCTGAGTTCTGGCGT SSGVVF GGTATTC

958 CGTWDYSL 1782 TGCGGAACATGGGATTACAGCCTGAGTGCCTGGGT SAWVF GTTC

959 CGTWDYSL 1783 TGCGGAACATGGGATTACAGTCTGAGTGCTGAGGT SAEVF GTTC

960 CGTWDYSL 1784 TGCGGAACATGGGATTACAGCCTGCGTCGTGCGAT RRAIF ATTC

961 CGTWDWS 1785 TGCGGAACATGGGATTGGAGCCTCATTCTTCAATT LILQLF GTTC

962 CGTWDVTL 1786 TGCGGAACATGGGATGTCACCTTGCATACTGGGGT HTGVF GTTC

963 CGTWDVTL 1787 TGCGGAACATGGGATGTCACCTTGCATATTGGGGT HIGVF GTTC

964 CGTWDVTL 1788 TGCGGAACATGGGATGTCACCTTGCATGCTGGGGT HAGVF GTTC

965 CGTWDVSL 1789 TGCGGAACATGGGATGTCAGTTTGTATAGTGGCGG YSGGVF GGTCTTC

966 CGTWDVSL 1790 TGTGGAACATGGGATGTCAGCCTGACTTCTTTCGTC TSFVF TTC

967 CGTWDVSL 1791 TGCGGAACATGGGATGTCAGCCTGAGTGTTGGGGT SVGVL GCTC

968 CGTWDVSL 1792 TGCGGAACGTGGGATGTCAGCCTGAGTGCTGGCGA SAGDVVF TGTAGTTTTC

969 CGTWDVSL 1793 TGCGGAACATGGGATGTCAGCCTGAATGTCGTGGT NVVVF TTTC

970 CGTWDVSL 1794 TGCGGAACATGGGATGTCAGCCTGAATACTCAGGT NTQVF GTTC

971 CGTWDVSL 1795 TGCGGCACATGGGATGTGAGCCTGGGTGCGCTGTT GALF C

972 CGTWDVN 1796 TGCGGAACGTGGGACGTTAATCTGAAAACTGTCGT LKTVVF TTTC

973 CGTWDVIL 1797 TGCGGAACATGGGATGTCATCCTGAGTGCTGAGGT SAEVF ATTC

974 CGTWDTTV 1798 TGCGGAACATGGGATACCACCGTGAGTGCTGTGGT SAVVF TTTC

975 CGTWDTTL 1799 TGCGGAACATGGGATACCACCCTGACTGCCTGGGT TAWVF GTTC

976 CGTWDTTL 1800 TGCGGAACATGGGACACCACCTTGAGTGTTTTCCT SVFLF ATTC

977 CGTWDTSV 1801 TGCGGGACTTGGGATACCAGTGTGAGTGCTGGGGT SAGVF GTTC 978 CGTWDTSV 1802 TGCGGAACATGGGATACCAGTGTGATTTCTTGGGT ISWVF TTTC

979 CGTWDTSR 1803 TGCGGAACATGGGATACCAGTCGGAGTTCTCTCTA SSLYVVF TGTGGTCTTC

980 CGTWDTSR 1804 TGCGGAACATGGGATACCAGCCGGAGTGCTTGGGT SAWVF ATTC

981 CGTWDTSR 1805 TGCGGAACATGGGATACCAGCCGGAATCCTGGAGG NPGGIF AATTTTC

982 CGTWDTSR 1806 TGCGGAACATGGGACACCAGTCGGGGTCATGTTTT GHVF C

983 CGTWDTSP 1807 TGCGGAACATGGGATACCAGCCCGAGTACTGGCCA STGQVLF GGTGCTTTTC

984 CGTWDTSP 1808 TGCGGAACATGGGATACCAGCCCGAGTGCCTGGGT SAWVF GTTC

985 CGTWDTSL 1809 TGCGGAACATGGGATACTAGCCTGACCTGGGTGTT TWVF C

986 CGTWDTSL 1810 TGCGGAACATGGGATACCAGCCTGACGTGGTTCGC TWFAVF AGTGTTC

987 CGTWDTSL 1811 TGCGGAACATGGGATACCAGCCTGACTGTTGTGGT TVVVF ATTC

988 CGTWDTSL 1812 TGCGGAACATGGGATACCAGCCTGACTACTTCTTG TTSWVF GGTGTTC

989 CGTWDTSL 1813 TGCGGAACATGGGATACCAGCCTGACCACTGGTCC TTGPFWVF TTTTTGGGTGTTC

990 CGTWDTSL 1814

TPFYVF GTCTTC

991 CGTWDTSL 1815 TGCGGAACATGGGATACCAGCCTGACTGCTTATGT TAYVF CTTC

992 CGTWDTSL 1816 TGCGGAACATGGGATACCAGCCTGACTGCTTGGGT TAWVF GTTC

993 CGTWDTSL 1817 TGCGGAACATGGGATACCAGCCTGACTGCGTGGGG TAWGVF GGTGTTC

994 CGTWDTSL 1818 TGCGGCACATGGGATACCAGCCTGACTGCGGTGGT TAVVL TCTC

995 CGTWDTSL 1819 TGCGGAACCTGGGATACCAGCCTGACTGCTCGGGT TARVF TTTC

996 CGTWDTSL 1820 TGCGGAACATGGGATACCAGCCTGACTGCGATTGT TAIVF CTTC

997 CGTWDTSL 1821 TGCGGAACATGGGATACCAGCCTGACTGCTGGTGT TAGVF CTTC

998 CGTWDTSL 1822 TGCGGAACATGGGATACCAGCCTGAGTGTTTATGT SVYVF CTTC

999 CGTWDTSL 1823 TGCGGAACATGGGATACCAGCCTGAGTGTGGTGTT SWF C

1000 CGTWDTSL 1824 TGCGGGACATGGGATACCAGCCTGAGTGTTGGGGA SVGEF ATTC

1001 CGTWDTSL 1825 TGCGGAACATGGGATACCAGCCTGAGTACTTGGGT STWVF GTTC

1002 CGTWDTSL 1826 TGCGGAACATGGGATACCAGCCTGAGTACTGTGGT STVVF ATTC 1003 CGTWDTSL 1827 TGCGGAACATGGGATACCAGCCTGAGTACTGGCCA STGQVLF GGTGCTTTTC

1004 CGTWDTSL 1828 TGCGGCACATGGGATACCAGCCTGAGCACTGGTCC STGPLWVF TCTTTGGGTGTTC

1005 CGTWDTSL 1829 TGCGGAACTTGGGATACCAGCCTGAGTTCTTATGT SSYVF CTTC

1006 CGTWDTSL 1830 TGCGGAACATGGGATACCAGCCTGAGTTCTGTGGT SSVVF CTTC

1007 CGTWDTSL 1831 TGCGGAACATGGGATACCAGCCTGAGTTCTAGATA SSRYIF CATATTC

1008 CGTWDTSL 1832 TGCGGAACATGGGATACCAGCCTGAGTTCTAGATT SSRFIF CATATTC

1009 CGTWDTSL 1833 TGCGGAACATGGGATACCAGCCTGAGTTCTGGGTG SSGWVF GGTGTTC

1010 CGTWDTSL 1834 TGCGGAACATGGGATACCAGCCTGAGTCGGTATGT SRYVF GTTC

1011 CGTWDTSL 1835 TGCGGAACTTGGGATACCAGTCTGAGTCAATGGCT SQWLF GTTC

1012 CGTWDTSL 1836 TGCGGAACATGGGATACCAGCCTGAGTCCTGGCCT SPGLWVF TTGGGTGTTC

1013 CGTWDTSL 1837 TGCGGAACATGGGATACCAGCCTGAGTAATTATGT SNYVF CTTC

1014 CGTWDTSL 1838 TGCGGAACATGGGATACCAGCCTAAGTATTTGGGT SIWVF GTTC

1015 CGTWDTSL 1839 TGCGGCACATGGGATACCAGCCTGAGCATTGGTCC SIGPFWVF TTTTTGGGTGTTC

1016 CGTWDTSL 1840 TGCGGAACATGGGATACCAGCCTGAGTGGTTGGGT SGWVF GTTC

1017 CGTWDTSL 1841 TGCGGAACATGGGATACCAGCCTGAGTGGTACAGT SGTVF GTTC

1018 CGTWDTSL 1842 TGCGGAACATGGGATACTAGTCTGAGTGGTGGCCA SGGQVF GGTGTTC

1019 CGTWDTSL 1843 TGCGGAACATGGGATACCAGCCTGAGTGGTGGGAT SGGIF ATTC

1020 CGTWDTSL 1844 TGCGGAACATGGGATACCAGCCTGAGTGGTGAGGA SGEDVVI TGTGGTAATC

1021 CGTWDTSL 1845 TGCGGAACATGGGATACCAGCCTGAGTTTCCTTTA SFLYAF TGCTTTC

1022 CGTWDTSL 1846 TGCGGAACATGGGATACCAGCCTGAGTGAGGTCGT SEWF ATTC

1023 CGTWDTSL 1847 TGCGGAACATGGGATACCAGCCTGAGTGAAGTGTT SEVF C

1024 CGTWDTSL 1848 TGCGGAACATGGGATACTAGCCTGAGTGAAAATTG SENWVF GGTGTTC

1025 CGTWDTSL 1849 TGCGGAACATGGGATACCAGCCTGAGTGCCTACAT SAYIF ATTC

1026 CGTWDTSL 1850 TGCGGAACATGGGATACCAGCCTGAGTGCTGTGGT SAVVL ACTC

1027 CGTWDTSL 1851 TGCGGAACATGGGATACCAGCCTGAGTGCTGTTTT SAVF C 1028 CGTWDTSL 1852 TGCGGAACATGGGATACCAGCCTGAGTGCCCGGGT SARVF GTTC

1029 CGTWDTSL 1853 TGCGGCACATGGGATACCAGCCTGAGTGCCCGCCA SARQVF GGTATTC

1030 CGTWDTSL 1854 TGCGGAACATGGGATACCAGCCTGAGTGCTTTGGT SALVF TTTC

1031 CGTWDTSL 1855 TGCGGAACATGGGATACCAGCCTGAGTGCTAAGGT SAKVF GTTC

1032 CGTWDTSL 1856 TGCGGAACATGGGATACCAGCCTGAGTGCGAAAAT SAKIF CTTC

1033 CGTWDTSL 1857 TGCGGAACATGGGATACCAGCCTGAGTGCCAAGGC SAKAVF GGTATTC

1034 CGTWDTSL 1858 TGCGGAACATGGGATACCAGCCTGAGTGCCCATGC SAHAVF TGTGTTC

1035 CGTWDTSL 1859 TGCGGAACATGGGATACCAGCCTGAGTGCTGGCTA SAGYVF TGTCTTC

1036 CGTWDTSL 1860 TGCGGAACATGGGACACCAGTCTGAGTGCTGGCCG SAGRWVF CTGGGTGTTC

1037 CGTWDTSL 1861 TGCGGAACATGGGATACCAGCCTGAGTGCTGGGAT SAGIF ATTC

1038 CGTWDTSL 1862 TGCGGAACATGGGATACCAGCCTGAGTGCTGGTGG SAGGFRVF GTTCCGGGTCTTC

1039 CGTWDTSL 1863 TGCGGAACATGGGATACCAGCCTGAGTGCTGGGGC SAGAF ATTC

1040 CGTWDTSL 1864 TGCGGAACATGGGATACCAGTCTGAGTGCTGATTG SADWFF GTTTTTC

1041 CGTWDTSL 1865 TGCGGAACATGGGATACCAGCCTGAGTGCTGATGA SADEYVF ATATGTCTTC

1042 CGTWDTSL 1866 TGCGGCACATGGGATACCAGCCTGAGTGCGGCTTG SAAWVF GGTGTTC

1043 CGTWDTSL 1867 TGCGGAACATGGGATACCAGCCTGAGTGCTGCGCT SAALF ATTC

1044 CGTWDTSL 1868 TGCGGAACATGGGATACCAGCCTGAGTGCTGCGGG SAAGVF GGTTTTC

1045 CGTWDTSL 1869 TGCGGAACATGGGATACCAGCCTGAGAGTTGTGGT RVVVF TTTC

1046 CGTWDTSL 1870 TGCGGAACATGGGATACCAGCCTGAGAACCTGGGT RTWVF ATTC

1047 CGTWDTSL 1871 TGCGGAACGTGGGATACCAGCCTGAGGGGTGCAGT RGAVF GTTC

1048 CGTWDTSL 1872 TGCGGAACATGGGATACCAGCCTGCGTGCTGTGGT RAVVF ATTC

1049 CGTWDTSL 1873 TGCGGAACATGGGATACAAGCCTGAATGTAGTTTA NVVYVF TGTCTTC

1050 CGTWDTSL 1874 TGCGGAACATGGGATACCAGCCTCAACACCTACCT NTYLF GTTC

1051 CGTWDTSL 1875 TGCGGAACATGGGATACTAGCCTGAACTTCGCTTG NFAWLF GCTGTTC

1052 CGTWDTSL 1876 TGCGGCACATGGGATACCAGCCTTCTTGTGTGGCTT LVWLF TTC 1053 CGTWDTSL 1877 TGCGGAACATGGGATACCAGTCTGAAGACGTGGGT KTWVF GTTC

1054 CGTWDTSL 1878 TGCGGAACATGGGATACCAGTCTGATTGTCTGGGT IVWVF GTTC

1055 CGTWDTSL 1879 TGCGGAACATGGGATACCAGCCTAATTACTGGGGT ITGVF GTTC

1056 CGTWDTSL 1880 TGCGGAACATGGGATACCAGCCTGATTAGCGTGGT ISVVF ATTC

1057 CGTWDTSL 1881 TGCGGAACATGGGATACCAGCCTGATTGCTTATGT IAYVF CTTC

1058 CGTWDTSL 1882 TGCGGAACATGGGATACCAGCCTGCACACTGAGTT HTELF GTTC

1059 CGTWDTSL 1883 TGCGGAACTTGGGATACCAGCCTGGGTTCTTATGT GSYVF CTTC

1060 CGTWDTSL 1884 TGCGGAACATGGGATACCAGCCTGGGTTCTCTTTG GSLWVF GGTGTTC

1061 CGTWDTSL 1885 TGCGGTACATGGGATACCAGCCTGGGTTCTGGGGT GSGVF ATTC

1062 CGTWDTSL 1886 TGCGGAACTTGGGATACCAGTCTGGGTGGTAGAGG GGRGVF GGTCTTC

1063 CGTWDTSL 1887 TGCGGAACATGGGATACCAGCCTGGGTGCTTGGGT GAWVF GTTC

1064 CGTWDTSL 1888 TGCGGAACATGGGATACCAGCCTGGGTGCCGTGGT GAVVF ATTC

1065 CGTWDTSL 1889 TGCGGAACATGGGATACCAGCCTGGGTGCTGGGGT GAGVF ATTC

1066 CGTWDTSL 1890 TGCGGAACATGGGATACCAGCCTGGGTGCTGGCCT GAGLF ATTC

1067 CGTWDTSL 1891 TGCGGAACATGGGATACCAGTCTGGATGCTGTGGT DAVVF TTTC

1068 CGTWDTSL 1892 TGCGGGACTTGGGATACCAGCCTGGATGCTGTGCT DAVLF GTTC

1069 CGTWDTSL 1893 TGCGGAACATGGGATACCAGCCTGGCTTGGGTGTT AWVF C

1070 CGTWDTSL 1894 TGCGGAACATGGGATACCAGCCTGGCGACTGGACT ATGLF GTTC

1071 CGTWDTSL 1895 TGCGGGACATGGGATACCAGCCTGGCCCCTGTAGT APVVF CTTC

1072 CGTWDTRL 1896 TGCGGAACATGGGACACCCGCCTGACTATTGTGAT TIVIF CTTC

1073 CGTWDTRL 1897 TGTGGAACATGGGACACCAGGCTGAGTGTTTGGCT SVWLF GTTC

1074 CGTWDTRL 1898 TGCGGAACGTGGGACACCAGACTGAGTGTTGGGGT SVGVF TTTC

1075 CGTWDTRL 1899 TGCGGCACATGGGATACCAGACTGAGTACTGTAAT STVIF TTTC

1076 CGTWDTRL 1900 TGCGGAACATGGGATACCCGCCTGAGTTCTGTGGT SSVVF CTTC

1077 CGTWDTRL 1901 TGCGGAACATGGGATACCCGCCTGAGTATTGTGGT SIVVF TTTC 1078 CGTWDTRL 1902 TGCGGAACATGGGATACCAGACTGAGTGCCTATGT SAYVVF GGTATTC

1079 CGTWDTRL 1903 TGCGGAACCTGGGACACCCGCCTGAGTGCGTGGGT SAWVF GTTC

1080 CGTWDTRL 1904 TGCGGAACATGGGATACCAGACTGAGTGCTGTGGT SAVVF GTTC

1081 CGTWDTRL 1905 TGCGGAACATGGGATACCCGCCTGAGTGCTGGGTT SAGLF GTTC

1082 CGTWDTRL 1906 TGCGGAACATGGGATACCAGACTGAGTGCTGGTGG SAGGVF GGTGTTC

1083 CGTWDTRL 1907 TGCGGAACATGGGATACCAGATTGAATGTGTGGCT NVWLF ATTC

1084 CGTWDTN 1908 TGCGGAACATGGGATACCAACCGGGAAGTTGTGCT REVVLL CCTC

1085 CGTWDTN L 1909 TGCGGAACATGGGATACCAACCTGCGTGCCCATGT RAHVF CTTC

1086 CGTWDTN L 1910 TGCGGAACATGGGATACTAATCTGCCCGCTGTAGT PAVVF GTTC

1087 CGTWDTN L 1911 TGCGGAACATGGGACACCAATTTGGGTGGGGTGTT GGVF C

1088 CGTWDTIV 1912 TGCGGAACATGGGATACCATCGTGAGTATTGGGGT SIGVF GTTC

1089 CGTWDTIL 1913 TGCGGAACATGGGATACCATCCTGAGTGCGGTGGT SAVVF GTTC

1090 CGTWDTIL 1914 TGCGGCACATGGGATACCATCCTGAGTGCTGAGGT SAEVF GTTC

1091 CGTWDTHL 1915 TGCGGAACATGGGATACCCACCTGGGTGTGGTTTT GVVF C

1092 CGTWDTGP 1916 TGCGGAACATGGGATACCGGCCCGAGCCCTCATTG SPHWLF GCTGTTC

1093 CGTWDTGL 1917 TGCGGAACATGGGATACCGGCCTGACTTTTGGAGG TFGGVF CGTGTTC

1094 CGTWDTGL 1918 TGCGGAACATGGGATACCGGCCTGACTGCTTTTGT TAFVF CTTC

1095 CGTWDTGL 1919 TGCGGAACATGGGATACCGGCCTGAGTGTTTGGGT SVWVF GTTC

1096 CGTWDTGL 1920 TGCGGAACATGGGATACCGGCCTGAGTACTGGGAT STGIF TTTC

1097 CGTWDTGL 1921 TGCGGAACATGGGATACCGGCCTGAGTTCCCTGCT SSLLF CTTC

1098 CGTWDTGL 1922 TGCGGAACGTGGGACACCGGCCTGAGTATTGTGGT SIVVF GTTC

1099 CGTWDTGL 1923 TGCGGAACGTGGGACACCGGCCTGAGTTTTGTGGT SFVVF GTTC

1100 CGTWDTGL 1924 TGCGGAACATGGGATACCGGCCTGAGTGCTTGGGT SAWVF GTTC

1101 CGTWDTGL 1925 TGCGGAACATGGGATACCGGCCTGAGTGCTGGTGT SAGVVF GGTATTC

1102 CGTWDTGL 1926 TGCGGAACATGGGATACCGGTCTGAGGGGTTGGAT RGWIF TTTC 1103 CGTWDTEL 1927 TGCGGAACATGGGATACCGAGCTAAGTGCGGGGGT SAGVF CTTC

1104 CGTWDTAL 1928 TGCGGAACGTGGGATACCGCCCTGACTGCTGGGGT TAGVF GTTC

1105 CGTWDTAL 1929 TGCGGAACATGGGATACTGCCCTGAGTCTTGTGGT SLWF CTTC

1106 CGTWDTAL 1930 TGCGGAACATGGGATACCGCCCTGAGTGCCTGGCT SAWLF GTTC

1107 CGTWDTAL 1931 TGCGGCACATGGGATACCGCCCTGAGTGCTGGGGT SAGVF GTTC

1108 CGTWDTAL 1932 TGCGGAACATGGGATACCGCCCTGCGTGGCGTGCT RGVLF GTTC

1109 CGTWDTAL 1933 TGCGGAACATGGGATACCGCCCTGAAAGAATGGCT KEWLF GTTC

1110 CGTWDRTL 1934 TGCGGAACATGGGATAGGACCCTGACTGCTGGCGA TAGDVLF TGTGCTCTTC

1111 CGTWDRSV 1935 TGCGGAACATGGGATAGAAGCGTGACTTATGTCTT TYVF C

1112 CGTWDRSR 1936 TGCGGAACATGGGATCGCAGCCGAAATGAATGGGT NEWVF GTTC

1113 CGTWDRSL 1937 TGCGGAACATGGGATCGCAGTCTGACTGTTTGGGT TVWVF CTTC

1114 CGTWDRSL 1938 TGCGGAACATGGGATCGCAGCCTGACTCCTGGGTG TPGWLF GTTGTTC

1115 CGTWDRSL 1939 TGCGGAACATGGGATAGAAGCCTGACTGCTTGGGT TAWVF GTTC

1116 CGTWDRSL 1940 TGCGGAACATGGGACCGCAGCCTGAGTGTTGTGGT SWVF ATTC

1117 CGTWDRSL 1941 TGCGGCACATGGGATCGCAGCCTGAGTGTAGTCTT SWF C

1118 CGTWDRSL 1942 TGCGGAACATGGGATAGGAGCCTGAGTGTTCAATT SVQLF GTTC

1119 CGTWDRSL 1943 TGCGGAACATGGGATCGCAGCCTCAGTGTTCTTTG SVLWVF GGTGTTC

1120 CGTWDRSL 1944 TGCGGAACATGGGATCGCAGCCTGAGTGTTGGATT SVGLF ATTC

1121 CGTWDRSL 1945 TGCGGAACATGGGATCGCAGCCTGAGTACTTGGGT STWVF GTTC

1122 CGTWDRSL 1946 TGCGGAACATGGGATAGAAGCCTGAGTACTCATTG STHWVL GGTGCTC

1123 CGTWDRSL 1947 TGCGGAACATGGGATAGAAGCCTGAGTACTCATTG STHWVF GGTGTTC

1124 CGTWDRSL 1948 TGCGGAACCTGGGATCGAAGCCTGAGTTCTGCGGT SSAVF GTTC

1125 CGTWDRSL 1949 TGCGGAACATGGGACAGAAGCCTGAGTCCCTCTTA SPSYVF TGTCTTC

1126 CGTWDRSL 1950 TGCGGAACATGGGATAGGAGCCTGAGTGGTGAGGT SGEVF GTTC

1127 CGTWDRSL 1951 TGCGGAACATGGGATAGGAGCCTGAGTGGTGCGGT SGAVF GTTC 1128 CGTWDRSL 1952 TGCGGAACATGGGATCGCAGCCTGAGTGCTGTGGC SAVAF ATTC

1129 CGTWDRSL 1953 TGCGGAACATGGGATAGGAGCCTGAGTGCCGGGG SAGGEF GGGAATTC

1130 CGTWDRSL 1954

SAFWVF GGTGTTC

1131 CGTWDRSL 1955 TGCGGAACATGGGATAGGAGCCTGAGTGCTGCGGT SAAVF GTTC

1132 CGTWDRSL 1956 TGCGGAACATGGGATAGGAGCCTGAGTGCTGCACT SAALF CTTC

1133 CGTWDRSL 1957 TGCGGAACATGGGATCGCAGCCTGAGAGTGTTC RVF

1134 CGTWDRSL 1958 TGCGGTACATGGGACAGAAGCCTTAATTGGGTGTT NWVF C

1135 CGTWDRSL 1959 TGCGGAACATGGGATCGCAGCCTGAATGTTTATGT NVYVF CTTC

1136 CGTWDRSL 1960 TGCGGAACATGGGATAGGAGCCTGAATGTTGGGGT NVGVF GTTC

1137 CGTWDRSL 1961 TGCGGAACATGGGATCGGAGCCTGCATGTGGTCTT HVVF C

1138 CGTWDRSL 1962 TGTGGAACATGGGATCGCAGCCTGGGTGGTTGGGT GGWVF GTTC

1139 CGTWDRSL 1963

GAFWVF GGTGTTC

1140 CGTWDRSL 1964 TGCGGAACATGGGATAGAAGCCTGTTTTGGGTGTT FWVF C

1141 CGTWDRSL 1965 TGCGGAACGTGGGATCGCAGCCTGGCTGCTGGGGT AAGVF GTTC

1142 CGTWDRRL 1966 TGCGGAACATGGGATAGGAGGTTGAGTGGTGTCGT SGVVF ATTC

1143 CGTWDRRL 1967 TGCGGAACGTGGGATCGCCGCCTAAGTGATGTGGT SDVVF ATTC

1144 CGTWDRRL 1968 TGCGGAACATGGGATAGGAGGCTGAGTGCTGTGGT SAVVF ATTC

1145 CGTWDRRL 1969 TGCGGAACATGGGATAGACGCCTGAATGTTGCGTT NVAFF CTTC

1146 CGTWDRRL 1970 TGTGGAACATGGGATAGGAGGCTGCTTGCTGTTTT LAVF C

1147 CGTWDRN 1971 TGCGGAACTTGGGATAGGAACCTGCGCGCCGTGGT LRAVVF CTTC

1148 CGTWDRLS 1972 TGCGGAACATGGGATAGGCTGAGTGCTGGGGTGTT AGVF C

1149 CGTWDRGP 1973 TGCGGAACATGGGATAGAGGCCCGAATACTGGGGT NTGVF ATTC

1150 CGTWDRG 1974 TGCGGAACATGGGATAGAGGCCTGAATACTGTTTA LNTVYVF CGTCTTC

1151 CGTWDNY 1975 TGCGGAACATGGGATAACTATGTGAGTGCCCCTTG VSAPWVF GGTGTTC

1152 CGTWDNY 1976 TGCGGAACATGGGATAACTACCTGAGTGCTGGCGA LSAGDVVF TGTGGTTTTC 1153 CGTWDNY 1977 TGCGGAACATGGGATAACTACCTGAGAGCTGGGGT LRAGVF CTTC

1154 CGTWDNY 1978 TGCGGAACATGGGACAATTATCTGGGTGCCGTGGT LGAVVF TTTC

1155 CGTWDNY 1979 TGCGGAACATGGGATAACTACCTGGGTGCGGGGGT LGAGVF GTTC

1156 CGTWDNT 1980 TGCGGAACATGGGATAACACCGTGAGTGCCCCTTG VSAPWVF GGTTTTC

1157 CGTWDNTL 1981 TGCGGAACATGGGATAACACCCTGAGTCTTTGGGT SLWVF GTTC

1158 CGTWDNTL 1982 TGCGGAACATGGGATAACACCCTGAGTGCTGGGGT SAGVF CTTC

1159 CGTWDNTL 1983 TGCGGAACATGGGACAACACTCTGCTTACTGTGTT LTVLF ATTC

1160 CGTWDNR 1984 TGCGGAACATGGGATAACAGACTGAGTAGTGTGAT LSSVIF TTTC

1161 CGTWDNR 1985 TGCGGAACATGGGATAACAGGTTGAGTGCTGTGGT LSAVVF CTTC

1162 CGTWDNR 1986 TGCGGAACATGGGATAACAGGCTGAGTGCTGGTGG LSAGGIF GATATTC

1163 CGTWDNR 1987 TGCGGAACATGGGATAACAGACTGAGTGCTGAGGT LSAEVF GTTC

1164 CGTWDNR 1988 TGTGGAACATGGGATAACAGACTGCGTGTTGGGGT LRVGVL TCTC

1165 CGTWDNR 1989 TGCGGAACATGGGATAATCGCCTGCTTGAGAATGT LLENVF CTTC

1166 CGTWDNN 1990 TGCGGAACATGGGATAACAACCTGCGTGCTGTCTT LRAVF C

1167 CGTWDNN 1991 TGCGGAACTTGGGATAATAACCTGCGTGCTGGAGT LRAGVF GTTC

1168 CGTWDNN 1992 TGCGGAACATGGGACAACAATTTGGGCGGTGGCCG LGGGRVF GGTGTTC

1169 CGTWDNN 1993 TGCGGAACATGGGATAACAACCTGGGTGCTGGCGT LGAGVL CCTC

1170 CGTWDNN 1994 TGCGGAACATGGGATAACAACCTGGGTGCTGGCGT LGAGVF CTTC

1171 CGTWDNIL 1995 TGCGGAACTTGGGATAACATCCTGAGCGCTGCGGT SAAVF GTTC

1172 CGTWDNIL 1996 TGCGGAACCTGGGATAACATCTTGGATGCAGGGGT DAGVF TTTC

1173 CGTWDND 1997 TGCGGAACATGGGATAACGACCTGAGTGGTTGGCT LSGWLF GTTC

1174 CGTWDND 1998 TGCGGAACATGGGATAACGACCTGAGTGCCTGGGT LSAWVF GTTC

1175 CGTWDLTL 1999 TGCGGAACATGGGATCTCACCCTGGGTGGTGTGGT GGVVF GTTC

1176 CGTWDLSL 2000 TGCGGAACATGGGATCTCAGCCTGAGTGCTGGGGT SAGVF ATTC

1177 CGTWDLSL 2001 TGCGGAACATGGGATCTCAGCCTGAAAGAATGGGT KEWVF GTTC 1178 CGTWDLSL 2002 TGCGGAACGTGGGATCTCAGCCTGGATGCTGTTGT DAVVF TTTC

1179 CGTWDLK 2003 TGCGGAACCTGGGACCTGAAGGTTTTC

VF

1180 CGTWDKTL 2004 TGCGGAACATGGGATAAGACTCTGAGTGTTTGGGT SVWVF GTTC

1181 CGTWDKSL 2005 TGCGGAACATGGGATAAGAGCCTGAGTGTTTGGGT SVWVF GTTC

1182 CGTWDKSL 2006 TGCGGAACATGGGATAAGAGCCTGAGTGGTGTGGT SGVVF ATTT

1183 CGTWDKSL 2007 TGCGGAACATGGGATAAGAGCCTGAGTGATTGGGT SDWVF GTTC

1184 CGTWDKSL 2008 TGCGGAACATGGGATAAGAGCCTGAGTGCTTTGGT SALVF TTTC

1185 CGTWDKSL 2009 TGCGGAACATGGGATAAGAGCCTGAGTGCTGGCGT SAGVF CTTC

1186 CGTWDKSL 2010 TGCGGAACATGGGATAAGAGCCTGAGTGCCGACGT SADVF CTTC

1187 CGTWDKR 2011 TGCGGAACATGGGATAAACGCCTGACTATTGTGGT LTIVVF CTTC

1188 CGTWDKR 2012 TGCGGAACATGGGATAAACGCCTGAGTGCCTGGGT LSAWVL GCTC

1189 CGTWDKN 2013 TGCGGAACATGGGATAAGAACCTGCGTGCTGTGGT LRAVVF CTTC

1190 CGTWDITL 2014 TGCGGAACATGGGATATCACCCTGAGTGGGTTTGT SGFVF CTTC

1191 CGTWDITL 2015 TGCGGAACATGGGATATCACCTTGCATACTGGAGT HTGVF ATTC

1192 CGTWDISV 2016 TGCGGAACATGGGATATCAGTGTGACTGTGGTGTT TVVF C

1193 CGTWDISV 2017 TGCGGAACATGGGATATCAGTGTGAGGGGTTATGC RGYAF CTTC

1194 CGTWDISR 2018 TGCGGAACATGGGATATCAGCCGTTGGGTTTTC WVF

1195 CGTWDISP 2019 TGCGGAACATGGGATATCAGCCCGAGTGCTTGGGT SAWVF GTTC

1196 CGTWDISL 2020 TGCGGAACATGGGATATTAGCCTGAGTGTCTGGGT SVWVF GTTC

1197 CGTWDISL 2021 TGCGGAACATGGGATATCAGCCTGAGTGTTGTATT SWF C

1198 CGTWDISL 2022 TGCGGAACTTGGGATATCAGCCTGAGTTCTGTGGT SSVVF GTTC

1199 CGTWDISL 2023 TGCGGAACATGGGATATCAGCCTGAGTCACTGGTT SHWLF GTTC

1200 CGTWDISL 2024 TGCGGAACATGGGATATCAGTCTGAGTGGTTGGGT SGWVF GTTC

1201 CGTWDISL 2025 TGCGGAACATGGGATATCAGCCTGAGTGGTCGAGT SGRVF GTTC

1202 CGTWDISL 2026 TGCGGAACATGGGACATCAGCCTGAGTGCTTGGGC SAWAF GTTC 1203 CGTWDISL 2027 TGCGGAACATGGGATATCAGCCTGAGTGCTGTGGT SAVVF TTTC

1204 CGTWDISL 2028 TGCGGGACATGGGACATCAGCCTGAGTGCTGTGAT SAVIF ATTC

1205 CGTWDISL 2029 TGCGGAACATGGGATATCAGCCTGAGTGCTGTGTT SAVF C

1206 CGTWDISL 2030 TGCGGAACATGGGATATCAGCCTGAGTGCCCGGGT SARVF GTTC

1207 CGTWDISL 2031 TGCGGAACATGGGATATCAGCCTGAGTGCCCTGGT SALVF GTTC

1208 CGTWDISL 2032 TGCGGAACATGGGATATTAGCCTGAGTGCCCATGT SAHVF CTTC

1209 CGTWDISL 2033 TGCGGAACATGGGATATCAGCCTGAGTGCTGGGGT SAGVVF GGTATTC

1210 CGTWDISL 2034 TGCGGAACATGGGATATCAGCCTGAGTGCCGGCCC SAGPYVF TTATGTCTTC

1211 CGTWDISL 2035 TGCGGCACATGGGATATCAGCCTGAGTGCTGGAGG SAGGVF GGTGTTC

1212 CGTWDISL 2036 TGCGGAACATGGGATATCAGCCTGAGTGCTGAGGT SAEVF TTTC

1213 CGTWDISL 2037 TGCGGAACATGGGATATCAGCCTGAGTGCTGCTGT SAAVF GTTC

1214 CGTWDISL 2038 TGCGGAACATGGGATATCAGCCTGCGTGCTGTGTT RAVF C

1215 CGTWDISL 2039 TGCGGAACATGGGATATTAGCCTGAATACTGGGGT NTGVF GTTC

1216 CGTWDISL 2040 TGCGGAACATGGGATATCAGCCTAAATAATTATGT N YVF CTTC

1217 CGTWDISLI 2041 TGCGGAACATGGGATATCAGCCTAATTGCTGGGGT AGVF ATTC

1218 CGTWDISL 2042 TGCGGAACATGGGATATCAGCCTGCATACTTGGCT HTWLF GTTC

1219 CGTWDIRL 2043 TGCGGAACATGGGATATCCGCCTGACCGATGAGCT TDELLF GTTATTC

1220 CGTWDIRL 2044 TGCGGAACATGGGATATCAGACTGAGCGGTTTTGT SGFVF TTTC

1221 CGTWDINL 2045 TGCGGAACATGGGATATCAACCTGGGTGCTGGGGG GAGGLYVF CCTTTATGTCTTC

1222 CGTWDIILS 2046 TGCGGAACATGGGATATCATCCTGAGTGCTGAGGT AEVF ATTC

1223 CGTWDHTL 2047 TGCGGAACATGGGATCACACCCTGAGTGCTGTCTT SAVF C

1224 CGTWDHTL 2048 TGCGGAACATGGGACCACACTCTGCTTACTGTGTT LTVLF ATTC

1225 CGTWDHSL 2049 TGCGGAACATGGGATCACAGCCTGACTGCTGTGGT TAVVF ATTC

1226 CGTWDHSL 2050 TGCGGAACCTGGGATCACAGCCTGACTGCTGGGAT TAGIF ATTC

1227 CGTWDHSL 2051 TGCGGAACATGGGATCACAGCCTGAGTGTTGTATT SWLF ATTC 1228 CGTWDHSL 2052 TGCGGAACATGGGATCACAGCCTGAGTTTGGTATT SLVF C

1229 CGTWDHSL 2053 TGCGGAACATGGGATCACAGCCTGTCTATTGGGGT SIGVF TTTC

1230 CGTWDHSL 2054 TGCGGAACATGGGATCACAGCCTGAGTGCTGGGGT SAGVF GTTC

1231 CGTWDHSL 2055 TGTGGAACTTGGGATCACAGCCTGAGTGCTTTCGT SAFVF GTTC

1232 CGTWDHSL 2056 TGCGGAACATGGGATCACAGTCTGAGTGCTGCTGT SAAVF TTTC

1233 CGTWDHN 2057 TGCGGAACATGGGACCACAATCTGCGTGCTGTCTT LRAVF C

1234 CGTWDFTL 2058 TGCGGGACATGGGATTTCACCCTGAGTGTTGGGCG SVGRF CTTC

1235 CGTWDFTL 2059 TGCGGAACATGGGATTTCACCCTGAGTGCTCCTGT SAPVF CTTC

1236 CGTWDFSV 2060 TGCGGAACGTGGGATTTCAGCGTGAGTGCTGGGTG SAGWVF GGTGTTC

1237 CGTWDFSL 2061 TGCGGAACGTGGGATTTCAGTCTTACTACCTGGTTA TTWLF TTC

1238 CGTWDFSL 2062 TGCGGAACATGGGATTTCAGCCTGAGTGTTTGGGT SVWVF GTTC

1239 CGTWDFSL 2063 TGCGGAACATGGGATTTCAGCCTGAGTACTGGGGT STGVF TTTC

1240 CGTWDFSL 2064 TGCGGCACATGGGATTTCAGCCTGAGTGGTGTGGT SGVVF ATTC

1241 CGTWDFSL 2065 TGCGGAACATGGGATTTCAGCCTGAGTGGTTTCGT SGFVF GTTC

1242 CGTWDFSL 2066 TGCGGAACATGGGATTTCAGCCTGAGTGCTGGGGT SAGVF GTTC

1243 CGTWDETV 2067 TGCGGAACATGGGATGAAACCGTGAGAGGTTGGGT RGWVF GTTC

1244 CGTWDESL 2068 TGCGGAACATGGGATGAAAGTCTGAGAAGCTGGGT RSWVF GTTC

1245 CGTWDER 2069 TGCGGAACTTGGGATGAGAGGCAGACTGATGAGTC QTDESYVF CTATGTCTTC

1246 CGTWDERL 2070 TGCGGAACATGGGATGAGAGACTCGTTGCTGGCCA VAGQVF GGTCTTC

1247 CGTWDERL 2071 TGCGGAACATGGGATGAGAGACTGAGTCCTGGAGC SPGAFF TTTTTTC

1248 CGTWDEK 2072 TGCGGAACATGGGATGAGAAGGTGTTC

VF

1249 CGTWDEG 2073 TGCGGAACCTGGGATGAAGGCCAGACTACTGATTT QTTDFFVF CTTTGTCTTC

1250 CGTWDDTL 2074 TGCGGAACATGGGATGACACCCTGGCTGGTGTGGT AGVVF CTTC

1251 CGTWDDR 2075 TGCGGAACATGGGATGACAGGCTGACTTCTGCGGT LTSAVF CTTC

1252 CGTWDDR 2076 TGCGGAACATGGGATGACAGACTGTTTGTTGTGGT LFWVF ATTC 1253 CGTWDDN 2077 TGCGGAACATGGGATGATAACCTGAGAGGTTGGGT LRGWVF GTTC

1254 CGTWDDN 2078 TGCGGAACATGGGATGACAACCTGCGTGGTGTCGT LRGVVF GTTC

1255 CGTWDDN 2079 TGCGGAACCTGGGATGACAATTTGAATATTGGAAG LNIGRVF GGTGTTC

1256 CGTWDDIL 2080 TGCGGAACATGGGATGACATCCTGAGTGCTGTGAT SAVIF ATTC

1257 CGTWDDIL 2081 TGCGGAACATGGGATGATATCCTGAGAGGTTGGGT RGWVF GTTC

1258 CGTWDATL 2082 TGCGGAACATGGGATGCCACCCTGAGTCCTGGGTG SPGWLF GTTATTC

1259 CGTWDAS 2083 TGCGGAACATGGGATGCCAGCGTGACTTCTTGGGT VTSWVF GTTC

1260 CGTWDASL 2084 TGCGGAACATGGGATGCCAGCCTGACTTCTGTGGT TSWF CTTC

1261 CGTWDASL 2085 TGCGGAACATGGGATGCCAGCCTGAGTGTTTGGGT SVWVF GTTC

1262 CGTWDASL 2086 TGCGGAACATGGGATGCCAGCCTGAGTGTTCCTTG SVPWVF GGTGTTC

1263 CGTWDASL 2087 TGCGGAACATGGGATGCCAGCCTGAGTGTGGCGGT SVAVF ATTC

1264 CGTWDASL 2088 TGCGGAACATGGGATGCCAGCCTGAGTACCTGGGT STWVF ATTC

1265 CGTWDASL 2089 TGCGGAACATGGGATGCCAGCCTGAGTGGTGTGGT SGVVF ATTC

1266 CGTWDASL 2090 TGCGGAACATGGGATGCCAGCCTGAGTGGTGGGGG SGGGEF AGAATTC

1267 CGTWDASL 2091 TGCGGAACATGGGATGCCAGCCTGAGTGCTGGGGT SAGVF GTTC

1268 CGTWDASL 2092 TGCGGAACATGGGATGCCAGCCTGAGTGCTGGGCT SAGLF TTTC

1269 CGTWDASL 2093 TGTGGCACATGGGATGCCAGCCTGAGTGCTGAAGT SAEVF CTTC

1270 CGTWDASL 2094 TGCGGAACATGGGATGCCAGCCTGAGTGCTGACTT SADFWVF TTGGGTGTTC

1271 CGTWDASL 2095 TGCGGAACATGGGATGCCAGCCTGAGAGTCTTCTT RVFF C

1272 CGTWDASL 2096 TGCGGAACATGGGATGCCAGTCTGAGGGCTGTGGT RAVVL ACTC

1273 CGTWDASL 2097 TGCGGAACATGGGATGCCAGCCTGAATATTTGGGT NIWVF TTTC

1274 CGTWDASL 2098 TGCGGGACATGGGATGCCAGCCTGAAGAATCTGGT KNLVF CTTC

1275 CGTWDASL 2099 TGCGGAACATGGGATGCCAGCCTGGGTGCCTGGGT GAWVF ATTC

1276 CGTWDASL 2100 TGCGGAACATGGGATGCCAGCCTGGGTGCTGTGGT GAVVF CTTC

1277 CGTWDASL 2101 TGCGGAACATGGGATGCCAGCCTGGGTGCGGGGGT GAGVF CTTC 1278 CGTWDAR 2102 TGCGGAACATGGGATGCTAGGCTGAGTGGCCTTTA LSGLYVF TGTCTTC

1279 CGTWDAR 2103 TGTGGAACCTGGGATGCGAGACTGGGTGGTGCAGT LGGAVF CTTC

1280 CGTWDAN 2104 TGCGGAACATGGGATGCCAATCTGCGTGCTGGGGT LRAGVF CTTC

1281 CGTWDAIIS 2105 TGCGGAACATGGGATGCTATCATAAGTGGTTGGGT GWVF GTTC

1282 CGTWDAG 2106 TGCGGAACATGGGATGCCGGCCAGAGTGTTTGGGT QSVWVF GTTC

1283 CGTWDAG 2107 TGCGGCACATGGGATGCCGGGCTGACTGGCCTTTA LTGLYVF TGTCTTC

1284 CGTWDAG 2108 TGCGGAACTTGGGATGCCGGTCTGAGTGTTTATGT LSVYVF CTTC

1285 CGTWDAG 2109 TGCGGGACATGGGATGCCGGCCTGAGTACTGGGGT LSTGVF CTTC

1286 CGTWDAG 2110 TGCGGAACATGGGATGCCGGCCTGAGTGGGGACGT LSGDVF TTTC

1287 CGTWDAG 2111 TGCGGAACATGGGATGCCGGCCTGAGTGCTGGTTA LSAGYVF TGTCTTC

1288 CGTWDAG 2112 TGCGGAACATGGGATGCCGGCCTGCGTGTTTGGGT LRVWVF GTTC

1289 CGTWDAG 2113 TGCGGAACATGGGATGCCGGCCTGAGGGAAATTTT LREIF C

1290 CGTWASSL 2114 TGCGGAACATGGGCCAGCAGCCTGAGTTCTTGGGT SSWVF GTTC

1291 CGTWAGSL 2115 TGCGGAACATGGGCTGGCAGCCTGAGTGGTCATGT SGHVF CTTC

1292 CGTWAGSL 2116 TGCGGAACATGGGCTGGCAGCCTGAGTGCCGCTTG SAAWVF GGTGTTC

1293 CGTWAGSL 2117 TGCGGAACATGGGCTGGCAGCCTGAATGTTTATTG NVYWVF GGTGTTC

1294 CGTWAGN 2118 TGCGGAACATGGGCTGGCAACCTGAGACCTAATTG LRPNWVF GGTGTTC

1295 CGTRGSLG 2119 TGCGGAACAAGGGGTAGCCTGGGTGGTGCGGTGTT GAVF C

1296 CGTRDTTL 2120 TGCGGAACAAGGGATACCACCCTGAGTGTCCCGGT SVPVF GTTC

1297 CGTRDTSL 2121 TGCGGAACACGGGATACCAGCCTCAATATTGAAAT NIEIF CTTC

1298 CGTRDTSL 2122 TGTGGAACACGGGATACCAGCCTGAATGATGTCTT NDVF C

1299 CGTRDTRL 2123 TGCGGAACACGGGATACCCGCCTGAGTATTGTGGT SIVVF TTTC

1300 CGTRDTILS 2124 TGCGGCACACGGGATACCATCCTGAGTGCTGAGGT AEVF GTTC

1301 CGTRDRSL 2125 TGCGGAACACGGGATAGAAGCCTGAGTGGTTGGGT SGWVF GTTC

1302 CGSWYYN 2126 TGCGGATCATGGTATTACAATGTCTTCCTTTTC VFLF 1303 CGSWHSSL 2127 TGCGGATCTTGGCATAGCAGCCTCAACCTTGTCGTC NLVVF TTC

1304 CGSWGSGL 2128 TGCGGATCATGGGGTAGTGGCCTGAGTGCCCCTTA SAPYVF TGTCTTC

1305 CGSWESGL 2129 TGCGGTTCGTGGGAAAGCGGCCTGGGTGCTTGGCT GAWLF GTTC

1306 CGSWDYG 2130 TGCGGATCCTGGGATTACGGCCTCCTACTCTTC LLLF

1307 CGSWDVSL 2131 TGCGGTTCATGGGATGTCAGCCTGACTGCTGTTTTC TAVF

1308 CGSWDVSL 2132 TGCGGATCCTGGGATGTCAGTCTCAATGTTGGCATT NVGIF TTC

1309 CGSWDTTL 2133 TGCGGATCATGGGATACCACCCTGCGTGCTTGGGT RAWVF GTTC

1310 CGSWDTSP 2134 TGCGGCTCGTGGGATACCAGCCCTGTCCGTGCTTG VRAWVF GGTGTTC

1311 CGSWDTSL 2135 TGCGGATCATGGGATACCAGCCTGAGTGTTTGGGT SVWVF GTTC

1312 CGSWDTSL 2136 TGCGGATCATGGGATACCAGCCTGAGTGCTGAGGT SAEVF GTTC

1313 CGSWDTSL 2137 TGCGGCTCGTGGGATACCAGCCTGCGTGCTTGGGT RAWVF GTTC

1314 CGSWDTSL 2138 TGCGGCTCGTGGGATACCAGCCTGCGTGCTTGGGC RAWAF GTTC

1315 CGSWDTSL 2139 TGCGGATCATGGGATACCAGCCTGGATGCTAGGCT DARLF GTTC

1316 CGSWDTIL 2140 TGCGGATCATGGGATACCATCCTGCTTGTCTATGTC LVYVF TTC

1317 CGSWDRW 2141 TGCGGATCATGGGATCGCTGGCAGGCTGCTGTCTT QAAVF C

1318 CGSWDRSL 2142 TGCGGATCATGGGATAGGAGCCTGAGTGGGTATGT SGYVF CTTC

1319 CGSWDRSL 2143 TGCGGATCATGGGATAGAAGCCTGAGTGCTTATGT SAYVF CTTC

1320 CGSWDRSL 2144 TGCGGATCATGGGATAGGAGCCTGAGTGCCGTGGT SAVVF TTTC

1321 CGSWDNTL 2145 TGCGGATCATGGGATAACACCTTGGGTGTTGTTCTC GVVLF TTC

1322 CGSWDNRL 2146 TGCGGATCGTGGGATAACAGACTAAGTACTGTCAT STVIF CTTC

1323 CGSWDNRL 2147 TGCGGAAGCTGGGATAATCGATTGAACACTGTGAT NTVIF TTTC

1324 CGSWDLSP 2148 TGCGGTTCATGGGATCTCAGCCCTGTACGTGTCCTT VRVLVF GTGTTC

1325 CGSWDLSL 2149 TGCGGATCATGGGATCTCAGCCTGAGTGCTGTCGT SAVVF TTTC

1326 CGSWDKN 2150 TGCGGATCATGGGATAAAAACCTGCGTGCTGTGCT LRAVLF GTTC

1327 CGSWDISL 2151 TGCGGCTCATGGGATATCAGCCTGAGTGCTGGGGT SAGVF GTTC 1328 CGSWDIRL 2152 TGCGGATCATGGGATATCAGACTGAGTGCAGAGGT SAEVF CTTC

1329 CGSWDIKL 2153 TGCGGATCATGGGACATCAAACTGAATATTGGGGT NIGVF ATTC

1330 CGSWDFSL 2154 TGCGGATCATGGGATTTCAGTCTCAATTATTTTGTC NYFVF TTC

1331 CGSWDASL 2155 TGCGGATCATGGGATGCCAGCCTGAGTACTGAGGT STEVF GTTC

1332 CGSWDAG 2156 TGCGGATCCTGGGATGCCGGCCTGCGTGGCTGGGT LRGWVF TTTC

1333 CGRWESSL 2157 TGCGGAAGATGGGAGAGCAGCCTGGGTGCTGTGGT GAVVF TTTC

1334 CGRWDFSL 2158 TGCGGAAGATGGGATTTTAGTCTGAGTGCTTATGT SAYVF CTTC

1335 CGQWDND 2159 TGCGGACAATGGGATAACGACCTGAGTGTTTGGGT LSVWVF GTTC

1336 CGPWHSSV 2160 TGCGGACCCTGGCATAGCAGCGTGACTAGTGGCCA TSGHVL CGTGCTC

1337 CGLWDASL 2161 TGCGGATTATGGGATGCCAGCCTGAGTGCTCCTAC SAPTWVF TTGGGTGTTC

1338 CGIWHTSL 2162 TGTGGAATATGGCACACTAGCCTGAGTGCTTGGGT SAWVF GTTC

1339 CGIWDYSL 2163 TGCGGAATATGGGATTACAGCCTGGATACTTGGGT DTWVF GTTC

1340 CGIWDTSL 2164 TGCGGCATATGGGATACCAGCCTGAGTGCTTGGGT SAWVF GTTC

1341 CGIWDTRL 2165 TGCGGAATTTGGGATACCAGGCTGAGTGTTTATGT

SVYVF CTTC

1342 CGIWDTRL 2166 TGCGGAATTTGGGATACCAGGCTGAGTGTTTATAT SVYIF CTTC

1343 CGIWDTNL 2167 TGTGGAATATGGGATACGAATCTGGGTTATCTCTTC GYLF

1344 CGIWDTGL 2168 TGCGGTATATGGGATACCGGCCTGAGTGCTGTGGT SAVVF ATTC

1345 CGIWDRSL 2169 TGCGGAATATGGGATCGCAGCCTGAGTGCTTGGGT SAWVF GTTT

1346 CGIRDTRLS 2170 TGCGGAATTCGGGATACCAGGCTGAGTGTTTATGT VYVF CTTC

1347 CGGWSSRL 2171 TGCGGAGGATGGAGTAGCAGACTGGGTGTTGGCCC GVGPVF AGTGTTT

1348 CGGWGSG 2172 TGCGGAGGATGGGGTAGCGGCCTGAGTGCTTGGGT LSAWVF GTTC

1349 CGGWDTSL 2173 TGCGGAGGATGGGATACCAGCCTGAGTGCTTGGGT SAWVF GTTC

1350 CGGWDRG 2174 TGCGGAGGATGGGATAGGGGCCTGGATGCTTGGGT LDAWVF TTTC

1351 CGAWRN 2175 TGCGGAGCATGGCGTAATAACGTGTGGGTGTTC VWVF

1352 CGAWNRR 2176 TGCGGAGCATGGAACAGGCGCCTGAATCCTCATTC LNPHSHWV TCATTGGGTGTTC F

1353 CGAWHNK 2177 TGCGGAGCCTGGCACAACAAACTGAGCGCGGTCTT LSAVF C

1354 CGAWGSSL 2178 TGCGGAGCATGGGGTAGCAGCCTGAGAGCTAGTGT RASVF CTTC

1355 CGAWGSG 2179 TGCGGAGCATGGGGTAGCGGCCTGAGTGCTTGGGT LSAWVF GTTC

1356 CGAWESSL 2180 TGCGGAGCATGGGAAAGTAGCCTGAGTGCCCCTTA SAPYVF TGTCTTC

1357 CGAWESSL 2181 TGCGGAGCATGGGAGAGCAGCCTCAATGTTGGACT NVGLI GATC

1358 CGAWESGR 2182 TGCGGAGCATGGGAGAGCGGCCGGAGTGCTGGGG SAGVVF TGGTGTTC

1359 CGAWDYS 2183 TGCGGAGCTTGGGATTACAGTGTGAGTGGTTGGGT VSGWVF GTTC

1360 CGAWDYS 2184 TGCGGAGCATGGGATTACAGCCTGACTGCCGGAGT LTAGVF ATTC

1361 CGAWDYR 2185 TGCGGAGCCTGGGATTACAGACTGAGTGCCGTGCT LSAVLF ATTC

1362 CGAWDVR 2186 TGCGGAGCGTGGGATGTTCGTCTGGATGTTGGGGT LDVGVF GTTC

1363 CGAWDTY 2187 TGCGGAGCATGGGATACCTACAGTTATGTCTTC SYVF

1364 CGAWDTTL 2188 TGCGGAGCATGGGATACGACCCTGAGTGGTGTGGT SGVVF ATTC

1365 CGAWDTTL 2189 TGCGGAGCGTGGGATACTACCCTGAGTGCTGTGAT SAVIF ATTC

1366 CGAWDTS 2190 TGCGGCGCATGGGATACCAGCCAGGGTGCGTCTTA QGASYVF TGTCTTT

1367 CGAWDTSP 2191 TGCGGAGCATGGGATACCAGCCCTGTACGTGCTGG VRAGVF GGTGTTC

1368 CGAWDTSL 2192 TGCGGAGCATGGGATACCAGCCTGTGGCTTTTC WLF

1369 CGAWDTSL 2193 TGCGGAGCATGGGATACCAGCCTGACTGTTTATGT TVYVF CTTC

1370 CGAWDTSL 2194 TGCGGAGCATGGGACACCAGTCTGACTGCTGGGGT TAGVF GTTC

1371 CGAWDTSL 2195 TGCGGAGCTTGGGATACCAGCCTGAGTACTGTGGT STVVF TTTC

1372 CGAWDTSL 2196 TGCGGAGCATGGGATACCAGCCTGAGTTCTAGATA SSRYIF CATATTC

1373 CGAWDTSL 2197 TGCGGAGCATGGGATACCAGCCTGAGTGGTTATGT SGYVF CTTC

1374 CGAWDTSL 2198 TGCGGAGCCTGGGATACCAGCCTGAGTGGCTGGGT SGWVF GTTC

1375 CGAWDTSL 2199 TGCGGAGCATGGGATACCAGTCTGAGTGGTGTGCT SGVLF ATTC

1376 CGAWDTSL 2200 TGCGGAGCTTGGGATACCAGCTTGAGTGGTCTTGT SGLVF TTTC

1377 CGAWDTSL 2201 TGCGGAGCTTGGGATACCAGCTTGAGTGGTTTTGTT SGFVF TTC

1378 CGAWDTSL 2202 TGCGGAGCATGGGATACCAGCCTGAGTGGTGAGGT SGEVF CTTT

1379 CGAWDTSL 2203 TGCGGAGCTTGGGATACCAGCTTGAGTGATTTTGTT SDFVF TTC

1380 CGAWDTSL 2204 TGCGGAGCATGGGATACCAGCCTGCGAACTGCGAT RTAIF ATTC

1381 CGAWDTSL 2205 TGCGGAGCATGGGATACCAGCCTGCGGCTTTTC RLF

1382 CGAWDTSL 2206 TGCGGAGCATGGGATACCAGCCTGAATGTTCATGT NVHVF CTTC

1383 CGAWDTSL 2207 TGCGGAGCATGGGATACCAGCCTCAATAAATGGGT NKWVF GTTC

1384 CGAWDTR 2208 TGCGGAGCATGGGATACCCGCCTCAGTGCGCGGCT LSARLF GTTC

1385 CGAWDTR 2209 TGCGGAGCATGGGATACCAGACTGAGGGGTTTTAT LRGFIF TTTC

1386 CGAWDTN 2210 TGCGGAGCATGGGATACTAATTTGGGGAATGTTCT LGNVLL CCTC

1387 CGAWDTN 2211 TGCGGGGCATGGGATACCAACCTGGGTAAATGGGT LGKWVF TTTC

1388 CGAWDTG 2212 TGCGGAGCATGGGATACCGGCCTTGAGTGGTATGT LEWYVF TTTT

1389 CGAWDRTS 2213 TGCGGAGCATGGGATAGGACTTCTGGATTGTGGCT GLWLF TTTC

1390 CGAWDRSL 2214 TGCGGAGCGTGGGATCGTAGCCTGGTTGCTGGACT VAGLF CTTC

1391 CGAWDRSL 2215 TGCGGAGCGTGGGATAGAAGCCTGACTGTTTATGT TVYVF CTTC

1392 CGAWDRSL 2216 TGCGGAGCATGGGATAGAAGCCTGAGTGGTTATGT SGYVF CTTC

1393 CGAWDRSL 2217 TGCGGAGCATGGGATAGAAGCCTGAGTGCTTATGT SAYVF CTTC

1394 CGAWDRSL 2218 TGCGGAGCATGGGATAGAAGCCTGAGTGCGGTGGT SAVVF ATTC

1395 CGAWDRSL 2219 TGCGGAGCATGGGATCGCAGCCTGAGTGCTGGGGT SAGVF TTTC

1396 CGAWDRSL 2220 TGCGGAGCGTGGGATCGCAGCCTGCGTATTGTGGT RIVVF ATTC

1397 CGAWDRSL 2221 TGCGGAGCATGGGATAGAAGTCTGAGGGCTTACGT RAYVF CTTC

1398 CGAWDRSL 2222 TGCGGAGCATGGGATAGAAGTCTGAATGTTTGGCT NVWLF GTTC

1399 CGAWDRG 2223 TGCGGCGCCTGGGATAGGGGCCTGAATGTCGGTTG LNVGWLF GCTTTTC

1400 CGAWDNR 2224 TGCGGCGCATGGGATAATAGACTGAGTATTTTGGC LSILAF CTTC

1401 CGAWDND 2225 TGCGGAGCTTGGGATAATGACCTGACAGCTTATGT LTAYVF CTTC

1402 CGAWDFSL 2226 TGCGGGGCATGGGATTTCAGCCTGACTCCTCTCTTC TPLF

1403 CGAWDDY 2227 TGCGGAGCCTGGGATGACTATCGGGGTGTGAGTAT RGVSIYVF TTATGTCTTC

1404 CGAWDDR 2228 TGTGGAGCATGGGATGACCGGCCTTCGAGTGCCGT PSSAVVF GGTTTTC

1405 CGAWDDR 2229 TGCGGAGCATGGGATGACAGACTGACTGTCGTTGT LTVVVF TTTC

1406 CGAWDDR 2230 TGCGGAGCGTGGGATGACAGGCTGGGTGCTGTGTT LGAVF C

1407 CGAWDAS 2231 TGCGGAGCGTGGGATGCCAGCCTGAATCCTGGCCG LNPGRAF GGCATTC

1408 CGAWDAG 2232 TGCGGAGCATGGGATGCCGGCCTGAGGGAAATTTT LREIF C

1409 CGAWAGSP 2233 TGCGGAGCTTGGGCTGGCAGTCCGAGTCCTTGGGT SPWVF TTTC

1410 CGAFDTTL 2234 TGCGGAGCATTCGACACCACCCTGAGTGCTGGCGT SAGVF TTTC

1411 CETWESSL 2235 TGCGAAACATGGGAGAGCAGCCTGAGTGTTGGGGT SVGVF CTTC

1412 CETWESSL 2236 TGCGAAACATGGGAAAGCAGCCTGAGGGTTTGGGT RVWVF GTTC

1413 CETWDTSL 2237 TGCGAAACGTGGGATACCAGCCTGAGTGGTGGGGT SGGVF GTTC

1414 CETWDTSL 2238 TGCGAAACATGGGATACCAGCCTGAGTGACTTTTA SDFYVF TGTCTTC

1415 CETWDTSL 2239 TGCGAAACATGGGATACCAGCCTGAGTGCCCTCTT SALF C

1416 CETWDTSL 2240 TGCGAAACATGGGATACCAGCCTGCGTGCTGAAGT RAEVF CTTC

1417 CETWDTSL 2241 TGCGAAACATGGGATACCAGCCTGAATGTTGTGGT NVVVF ATTC

1418 CETWDTSL 2242 TGCGAAACATGGGATACCAGCCTGGGTGCCGTGGT GAVVF GTTC

1419 CETWDRSL 2243 TGCGAAACATGGGATAGAAGCCTGAGTGGTGTGGT SGVVF ATTC

1420 CETWDRSL 2244 TGCGAAACATGGGATAGGAGCCTGAGTGCTTGGGT SAWVF GTTT

1421 CETWDRSL 2245 TGCGAAACATGGGATCGCAGCCTGAGTGCTGTGGT SAVVF CTTC

1422 CETWDRGL 2246 TGCGAGACGTGGGATAGAGGCCTGAGTGTTGTGGT SWVF TTTC

1423 CETWDRGL 2247 TGCGAAACATGGGATAGGGGCCTGAGTGCAGTGGT SAVVF ATTC

1424 CETWDHTL 2248 TGCGAAACATGGGATCACACCCTGAGTGTTGTGAT SWIF ATTC

1425 CETWDASL 2249 TGCGAAACATGGGATGCCAGCCTGACTGTTGTGTT TVVLF ATTC

1426 CETWDASL 2250 TGCGAAACATGGGATGCCAGCCTGAGTGCTGGGGT SAGVF GTTC

1427 CETWDAGL 2251 TGCGAAACGTGGGATGCCGGCCTGAGTGAGGTGGT SEWF GTTC

1428 CETFDTSLS 2252 TGCGAAACATTTGATACCAGCCTGAGTGTTGTAGT VVVF CTTC

1429 CETFDTSL 2253 TGCGAAACATTTGATACCAGCCTAAATATTGTAGT NIVVF CTTT

1430 CESWDRSR 2254 TGCGAATCATGGGATAGAAGCCGGATTGGTGTGGT IGVVF CTTC

1431 CESWDRSL 2255 TGCGAAAGTTGGGACAGGAGTCTGAGTGCCCGGGT SARVY GTAC

1432 CESWDRSL 2256 TGCGAATCCTGGGATAGGAGCCTGCGTGCCGTGGT RAVVF CTTC

1433 CESWDRSL 2257 TGCGAATCTTGGGATCGTAGTTTGATTGTGGTGTTC IVVF

1434 CESWDN L 2258 TGCGAAAGTTGGGATAACAATTTAAATGAGGTGGT NEVVF TTTC

1435 CEIWESSPS 2259 TGCGAAATATGGGAGAGCAGCCCGAGTGCTGACG ADDLVF ATTTGGTGTTC

1436 CEAWDTSL 2260 TGCGAAGCATGGGATACCAGCCTGAGTGGTGCGGT SGAVF GTTC

1437 CEAWDTSL 2261 TGCGAAGCATGGGATACCAGCCTGAGTGCCGGGGT SAGVF GTTC

1438 CEAWDTSL 2262 TGCGAAGCATGGGATACCAGCCTGGGTGGTGGGGT GGGVF GTTC

1439 CEAWDRSL 2263 TGCGAAGCATGGGATCGCAGCCTGACTGGTAGCCT TGSLF GTTC

1440 CEAWDRG 2264 TGCGAAGCGTGGGATAGGGGCCTGAGTGCAGTGGT LSAVVF ATTC

1441 CEAWDNIL 2265 TGCGAAGCCTGGGATAACATCCTGAGTACTGTGGT STVVF GTTC

1442 CEAWDISL 2266 TGCGAAGCATGGGACATCAGCCTGAGTGCTGGGGT SAGVF GTTC

1443 CEAWDAD 2267 TGCGAAGCATGGGATGCCGACCTGAGTGGTGCGGT LSGAVF GTTC

1444 CATWTGSF 2268 TGCGCAACATGGACTGGTAGTTTCAGAACTGGCCA RTGHYVF TTATGTCTTC

1445 CATWSSSP 2269 TGCGCAACATGGAGTAGCAGTCCCAGGGGGTGGGT RGWVF GTTC

1446 CATWHYSL 2270 TGCGCAACATGGCATTACAGCCTGAGTGCTGGCCG SAGRVF AGTGTTC

1447 CATWHTSL 2271 TGCGCAACATGGCATACCAGCCTGAGTATTGTGCA SIVQF GTTC

1448 CATWHSTL 2272 TGCGCAACATGGCATAGCACCCTGAGTGCTGATGT SADVLF GCTTTTC

1449 CATWHSSL 2273 TGCGCAACATGGCATAGCAGCCTGAGTGCTGGCCG SAGRLF ACTCTTC

1450 CATWHIAR 2274 TGCGCAACATGGCATATCGCTCGGAGTGCCTGGGT SAWVF GTTC

1451 CATWGSSQ 2275 TGCGCAACATGGGGTAGTAGTCAGAGTGCCGTGGT SAVVF ATTC

1452 CATWGSSL 2276 TGCGCAACATGGGGTAGCAGCCTGAGTGCTGGGGG SAGGVF TGTTTTC

1453 CATWEYSL 2277 TGTGCAACATGGGAATACAGCCTGAGTGTTGTGCT SWLF GTTC

1454 CATWETTR 2278 TGCGCAACATGGGAGACCACCCGACGTGCCTCTTT RASFVF TGTCTTC

1455 CATWETSL 2279 TGCGCAACATGGGAGACCAGCCTGAATGTTTATGT NVYVF CTTC

1456 CATWETSL 2280 TGCGCAACATGGGAAACTAGCCTGAATGTTGTGGT NVVVF CTTC

1457 CATWETSL 2281 TGCGCAACATGGGAGACCAGCCTGAATCTTTATGT NLYVF CTTC

1458 CATWETGL 2282 TGCGCAACATGGGAGACTGGCCTAAGTGCTGGAGA SAGEVF GGTGTTC

1459 CATWESTL 2283 TGCGCGACGTGGGAGAGTACCCTAAGTGTTGTGGT SWVF TTTC

1460 CATWESSL 2284

SIFVF CTTC

1461 CATWESSL 2285 TGCGCAACATGGGAAAGCAGCCTCAACACTTTTTA NTFYVF TGTCTTC

1462 CATWESRV 2286 TGCGCAACATGGGAGAGTAGGGTGGATACTCGAG DTRGLLF GGTTGTTATTC

1463 CATWESGL 2287 TGCGCAACATGGGAGAGCGGCCTGAGTGGTGCGG SGAGVF GGGTGTTC

1464 CATWEGSL 2288 TGCGCAACATGGGAAGGCAGCCTCAACACTTTTTA NTFYVF TGTCTTC

1465 CATWDYSL 2289 TGCGCAACTTGGGATTATAGCCTGAGTGCTGTGGT SAVVF GTTC

1466 CATWDYR 2290 TGCGCAACATGGGATTACAGACTGAGTATTGTGGT LSIVVF ATTC

1467 CATWDYN 2291 TGCGCAACATGGGATTATAACCTGGGAGCTGCGGT LGAAVF GTTC

1468 CATWDVTL 2292 TGCGCCACATGGGATGTCACCCTGGGTGTCTTGCA GVLHF TTTC

1469 CATWDTTL 2293 TGCGCAACATGGGATACAACACTGAGTGTCTGGGT SVWVF CTTC

1470 CATWDTTL 2294 TGCGCAACATGGGATACCACCCTGAGTGTAGTACT SWLF TTTC

1471 CATWDTTL 2295 TGCGCAACATGGGATACCACCCTGAGTGTTGAGGT SVEVF CTTC

1472 CATWDTSP 2296 TGCGCAACATGGGATACCAGCCCCAGCCTGAGTGG SLSGFWVF TTTTTGGGTGTTC

1473 CATWDTSL 2297 TGCGCAACATGGGATACCAGCCTGACTGGTGTGGT TGVVF ATTC

1474 CATWDTSL 2298 TGCGCAACATGGGATACCAGCCTGACTGGTGCGGT TGAVF GTTC

1475 CATWDTSL 2299 TGCGCAACATGGGATACCAGCCTGACTGCCTGGGT TAWVF ATTC

1476 CATWDTSL 2300 TGCGCAACATGGGATACCAGCCTGACTGCTGTGGT TAVVF TTTC

1477 CATWDTSL 2301 TGCGCAACATGGGATACTAGCCTGACTGCTAAGGT TAKVF GTTC

1478 CATWDTSL 2302 TGCGCAACATGGGACACCAGCCTGAGTGTTGTGGT SWVF TTTC

1479 CATWDTSL 2303 TGCGCTACTTGGGATACCAGCCTGAGTGTTGGGGT SVGVF ATTT

1480 CATWDTSL 2304 TGCGCAACATGGGATACCAGCCTGAGTTCTTGGGT SSWVF GTTC

1481 CATWDTSL 2305 TGCGCAACATGGGATACCAGCCTGAGTGGTGGGGT SGGVL ACTC

1482 CATWDTSL 2306 TGCGCAACATGGGATACCAGCCTGAGTGGTGGGGT SGGVF GTTC

1483 CATWDTSL 2307 TGCGCAACATGGGATACCAGCCTGAGTGGTGGCCG SGGRVF AGTGTTC

1484 CATWDTSL 2308 TGCGCAACATGGGATACCAGCCTGAGTGGTGACCG SGDRVF AGTGTTC

1485 CATWDTSL 2309 TGCGCAACGTGGGATACTAGCCTGAGTGAAGGGGT SEGVF GTTC

1486 CATWDTSL 2310 TGCGCAACCTGGGATACCAGCCTGAGTGCCGTGGT SAVVL GCTC

1487 CATWDTSL 2311 TGCGCAACATGGGATACCAGCCTGAGTGCTGTCTT SAVF C

1488 CATWDTSL 2312 TGCGCGACATGGGATACCAGCCTGAGTGCTCGGGT SARVF GTTC

1489 CATWDTSL 2313 TGCGCAACATGGGATACCAGCCTGAGTGCCTTATT SALF C

1490 CATWDTSL 2314 TGCGCAACATGGGATACCAGCCTGAGTGCTCATGT SAHVF CTTC

1491 CATWDTSL 2315 TGCGCAACATGGGATACCAGCCTGAGTGCTGGCCG SAGRVF GGTGTTC

1492 CATWDTSL 2316 TGCGCAACATGGGATACCAGCCTGAGTGCGGAGGT SAEVF CTTC

1493 CATWDTSL 2317 TGCGCAACATGGGATACCAGCCTGAGTGCTGATGC SADAGGGV TGGTGGGGGGGTCTTC

F

1494 CATWDTSL 2318 TGCGCAACATGGGATACCAGCCTGCGTGTCGTGGT RVVVF ATTC

1495 CATWDTSL 2319 TGCGCAACATGGGATACCAGCCTGAGAGGGGTGTT RGVF C

1496 CATWDTSL 2320 TGCGCAACATGGGATACCAGCCTGCCTGCGTGGGT PAWVF GTTC

1497 CATWDTSL 2321 TGTGCAACATGGGATACCAGCCTGAATGTTGGGGT NVGVF ATTC

1498 CATWDTSL 2322 TGCGCAACATGGGATACCAGCCTGGGTATTGTGTT GIVLF ATTT

1499 CATWDTSL 2323 TGCGCAACATGGGACACCAGCCTGGGTGCGCGTGT GARVVF GGTCTTC

1500 CATWDTSL 2324 TGTGCAACGTGGGATACCAGTCTAGGTGCCTTGTT GALF C

1501 CATWDTSL 2325 TGCGCAACATGGGATACCAGCCTGGCGACTGGACT ATGLF GTTC 1502 CATWDTSL 2326 TGCGCAACATGGGATACCAGCCTGGCTGCCTGGGT AAWVF ATTC

1503 CATWDTRL 2327 TGCGCAACCTGGGATACCAGGCTGAGTGCTGTGGT SAVVF CTTC

1504 CATWDTRL 2328 TGCGCAACATGGGATACCAGGCTGAGTGCTGGGGT SAGVF GTTC

1505 CATWDTRL 2329 TGTGCAACGTGGGACACACGTCTACTTATTACGGT LITVF TTTC

1506 CATWDTLL 2330 TGCGCAACATGGGACACCCTCCTGAGTGTTGAACT SVELF CTTC

1507 CATWDTG 2331 TGCGCAACATGGGATACTGGCCGCAATCCTCATGT RNPHVVF GGTCTTC

1508 CATWDTGL 2332 TGCGCAACATGGGATACCGGCCTGTCTTCGGTGTT SSVLF GTTC

1509 CATWDTGL 2333 TGCGCAACGTGGGATACCGGCCTGAGTGCGGTTTT SAVF C

1510 CATWDRTL 2334 TGCGCTACGTGGGATAGGACCCTGAGTATTGGAGT SIGVF CTTC

1511 CATWDRSV 2335 TGCGCAACGTGGGATCGCAGTGTGACTGCTGTGCT TAVLF CTTC

1512 CATWDRSL 2336 TGCGCAACCTGGGATAGGAGCCTGAGTGGTGTGGT SGVVF GTTC

1513 CATWDRSL 2337 TGCGCAACATGGGATAGAAGCCTGAGTGCTGTGGT SAVVF CTTC

1514 CATWDRSL 2338 TGCGCAACATGGGATAGAAGCCTGAGTGCTGTTCC SAVPWVF TTGGGTGTTC

1515 CATWDRSL 2339 TGCGCAACATGGGATCGCAGCCTGAGTGCTGGGGT SAGVF GTTC

1516 CATWDRSL 2340 TGCGCAACGTGGGATAGGAGCCTGCGTGCTGGGGT RAGVF GTTC

1517 CATWDRSL 2341 TGCGCAACATGGGATCGCAGTCTGAATGTTTATGT NVYVL CCTC

1518 CATWDRIL 2342 TGCGCAACGTGGGATCGCATCCTGAGCGCTGAGGT SAEVF GTTC

1519 CATWDRG 2343 TGCGCAACGTGGGATAGAGGCCTGAGTACTGGGGT LSTGVF GTTC

1520 CATWDNY 2344 TGCGCAACATGGGATAACTACCTGGGTGCTGCCGT LGAAVF GTTC

1521 CATWDNTP 2345 TGCGCAACATGGGATAACACGCCTTCGAATATTGT SNIVVF GGTATTC

1522 CATWDNTL 2346 TGCGCAACATGGGATAATACACTGAGTGTGTGGGT SVWVF CTTC

1523 CATWDNTL 2347 TGCGCAACATGGGATAACACCCTGAGTGTCAATTG SVNWVF GGTGTTC

1524 CATWDNTL 2348 TGCGCAACCTGGGATAACACACTGAATGTCTTTTA NVFYVF TGTTTTC

1525 CATWDNR 2349 TGTGCGACATGGGATAATCGGCTCAGTTCTGTGGT LSSVVF CTTC

1526 CATWDNR 2350 TGCGCAACATGGGATAACCGCCTGAGTGCTGGGGT LSAGVL GCTC 1552 CATWDATL 2376 TGCGCAACATGGGATGCGACCCTGAATACTGGGGT NTGVF GTTC

1553 CATWDASL 2377 TGCGCAACATGGGATGCCAGCCTGAGTGTTTGGCT SVWLL GCTC

1554 CATWDASL 2378 TGCGCGACATGGGATGCCAGCCTGAGTGGTGGGGT SGGVF GTTC

1555 CATRDTTL 2379 TGCGCAACACGGGATACCACCCTCAGCGCCGTTCT SAVLF GTTC

1556 CATLGSSLS 2380 TGCGCTACATTGGGTAGTAGCCTGAGTCTCTGGGT LWVF GTTC

1557 CATIETSLP 2381 TGCGCAACAATCGAAACTAGCCTGCCTGCCTGGGT AWVF ATTC

1558 CATGDRSL 2382 TGCGCAACAGGGGACAGAAGCCTGACTGTTGAGGT TVEVF ATTC

1559 CATGDLGL 2383 TGCGCTACAGGGGATCTCGGCCTGACCATAGTCTT TIVF C

1560 CASWDYR 2384 TGCGCATCATGGGATTACAGGGGGAGATCTGGTTG GRSGWVF GGTGTTC

1561 CASWDTTL 2385 TGCGCATCATGGGATACCACCCTGAATGTTGGGGT NVGVF GTTC

1562 CASWDTTL 2386 TGCGCTTCATGGGATACCACCCTGGGTTTTGTGTTA GFVLF TTC

1563 CASWDTSL 2387 TGCGCATCATGGGATACCAGCCTGAGTGGTGGTTA SGGYVF TGTCTTC

1564 CASWDTSL 2388 TGCGCATCATGGGATACCAGCCTCCGTGCTGGGGT RAGVF GTTC

1565 CASWDTSL 2389 TGCGCATCATGGGATACCAGCCTGGGTGCTGGGGT GAGVF GTTC

1566 CASWDRGL 2390 TGCGCATCATGGGACAGAGGCCTGAGTGCAGTGGT SAVVF GTTC

1567 CASWDNV 2391 TGTGCTAGTTGGGATAACGTCCTGCGTGGTGTGGT LRGVVF ATTC

1568 CASWDNRL 2392 TGCGCGTCATGGGATAACAGGCTGACTGCCGTGGT TAVVF TTTC

1569 CASWDASL 2393 TGCGCATCATGGGATGCAAGCCTGTCCGTCGCTTTC SVAF

1570 CASWDAG 2394 TGCGCTTCGTGGGATGCCGGCCTGAGTTCTTATGTC LSSYVF TTC

1571 CASGDTSL 2395 TGCGCATCCGGGGATACCAGCCTGAGTGGTGTGAT SGVIF ATTC

1572 CARWHTSL 2396 TGCGCAAGATGGCATACGAGCCTAAGTATTTGGGT SIWVF CTTC

1573 CAIWDTGL 2397 TGCGCAATATGGGATACCGGCCTGAGTCCTGGCCA SPGQVAF AGTTGCCTTC

1574 CAAWHSG 2398 TGCGCAGCATGGCATAGCGGCCTGGGTCTCCCGGT LGLPVF CTTC

1575 CAAWDYS 2399 TGCGCAGCATGGGATTACAGCCTGAGTGCTGGGGT LSAGVF GTTC

1576 CAAWDTTL 2400 TGCGCAGCCTGGGATACTACCCTGCGTGTTAGGCT RVRLF GTTC 1577 CAAWDTSL 2401 TGCGCAGCATGGGATACCAGCCTGACTGCCTGGGT TAWVF TTTC

1578 CAAWDTSL 2402 TGCGCAGCATGGGATACCAGCTTGAGTGGTGGGGT SGGVF GTTC

1579 CAAWDTSL 2403 TGCGCAGCATGGGATACCAGCCTGAGTGGCGAGGC SGEAVF TGTGTTC

1580 CAAWDTSL 2404 TGCGCAGCATGGGATACCAGCTTGAGTGGTGCGGT SGAVF GTTC

1581 CAAWDTSL 2405 TGCGCAGCATGGGATACCAGCCTGAGTGCCTGGGT SAWVF GTTC

1582 CAAWDTSL 2406 TGCGCAGCATGGGATACCAGCCTGAGTGCTGGGGT SAGVF ATTC

1583 CAAWDTSL 2407 TGCGCAGCATGGGATACCAGCCTGGATACTTATGT DTYVF CTTC

1584 CAAWDTR 2408 TGCGCTGCATGGGATACCCGTCTGAGTGGTGTGTT LSGVLF ATTC

1585 CAAWDTR 2409 TGCGCAGCATGGGATACCAGGCTGAGTGCTGGGGT LSAGVF GTTC

1586 CAAWDRSL 2410 TGCGCAGCATGGGATCGCAGTCTGAGTACTGGAGT STGVF TTTC

1587 CAAWDIRR 2411 TGCGCAGCGTGGGATATCCGCCGGTCTGTCCTTTTC SVLF

1588 CAAWDHT 2412 TGCGCTGCGTGGGATCACACTCAGCGTCTTTCCTTC QRLSF

1589 CAAWDHS 2413 TGCGCAGCATGGGATCACAGCCTGAGTGCTGGCCA LSAGQVF GGTGTTC

1590 CAAVDTGL 2414 TGCGCAGCAGTCGATACTGGTCTGAAAGAATGGGT KEWVF GTTC

[00210] The CDRs were prescreened to contain no amino acid liabilities, cryptic splice sites or nucleotide restriction sites. The CDR variation was observed in at least two individuals and comprises the near-germline space of single, double and triple mutations. The order of assembly is seen in FIG. 21C.

[00211] The VH domains that were designed include IGHV1-69 and IGHV3-30. Each of two heavy chain VH domains are assembled with their respective invariant 4 framework elements (FW1, FW2, FW3, FW4) and variable 3 CDR (HI, H2, H3) elements. For IGHV1-69, 417 variants were designed for HI and 258 variants were designed for H2. For IGHV3-30, 535 variants were designed for HI and 165 variants were designed for H2. For the CDR H3, the same cassette was used in both IGHVl-69 and IGHV-30 since both designed use an identical FW4, and because the edge of FW3 is also identical for both IGHVl-69 and IGHV3-30. The CDR H3 comprises an N-terminus and C-terminus element that are combinatorially joined to a central middle element to generate 1 x 10 ¹⁰ diversity. The N-terminal and middle element overlap with a "GGG" glycine codon. The middle and C-terminal element overlap with a "GGT" glycine codon. The CDR H3 comprises 5 subpools that were assembled separately. The various N-terminus and C-terminus elements comprise sequences as seen in Table 14.

Table 14. Sequences for N-terminus and C-terminus elements

[00212] Example 12. Enrichment for GPCR GLP1R Binding Proteins

[00213] Antibodies having CDR-H3 regions with a variant fragments of GPCR binding protein were generated by methods described herein were panned using cell-based methods to identified variants which are enriched for binding to particular GPCRs, as described in Example

10.

[00214] Variants of the GLP C-terminus peptide were identified (listed in Table 15) that when embedded in the CDR-H3 region of an antibody, were repeatedly and selectively enriched for binding to GPCR GLP1R.

Table 15. Sequences of GLP1 embedded in CDR-H3

2435 CAKHMSMQEGA VTGEGQ AAKEFI AWLVKGGLTYD S SED SGGAFDIW

2436 CAKHMSMQDYLVIGEGQAAKEFIAWLVKGRVRADLVGDAFDVW

[00215] While preferred embodiments of the present disclosure have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the disclosure. It should be understood that various alternatives to the embodiments of the disclosure described herein may be employed in practicing the disclosure. It is intended that the following claims define the scope of the disclosure and that methods and structures within the scope of these claims and their equivalents be covered thereby.