Title:
MOLECULAR ENGINEERING METHOD FOR IN VITRO EVOLUTION OF MEMBRANE PROTEIN
Document Type and Number:
WIPO Patent Application WO/2014/002424
Kind Code:
A1
Abstract:
The present invention addresses the problem of improving the efficiency of screening/selection of a membrane protein during molecular evolutionary engineering (in an enzyme evolution method, for example). The abovementioned problem was solved by providing a unilamellar liposome that comprises: (a) DNA that includes a promoter sequence, a translation initiation sequence, and a sequence that encodes a membrane protein; (b) RNA polymerase; (c) a ribonucleotide; and (d) a cell-free protein synthesis system. In one embodiment of the present invention, the membrane protein is a transporter and the unilamellar liposome additionally includes (e) a factor that binds to a ligand that is transported by the membrane protein.
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Inventors:
YOMO TETSUYA (JP)
MATSUURA TOMOAKI (JP)
SOGA HARUKA (JP)
WATANABE HAJIME (JP)
FUJII SATOSHI (JP)
MATSUURA TOMOAKI (JP)
SOGA HARUKA (JP)
WATANABE HAJIME (JP)
FUJII SATOSHI (JP)
Application Number:
PCT/JP2013/003767
Publication Date:
January 03, 2014
Filing Date:
June 17, 2013
Export Citation:
Assignee:
JAPAN SCIENCE & TECH AGENCY (JP)
International Classes:
C12P21/02; C12N15/09; C12Q1/68
Foreign References:
| JP2012210170A | 2012-11-01 |
Other References:
TAKEHIRO NISHIKAWA ET AL.: "Tanso Maku Liposome o Hannoba to suru Idenshi Screening System no Teiryoteki Hyoka", SYMPOSIUM ON MACROMOLECULES YOKOSHU, vol. 60, no. 2, 13 September 2011 (2011-09-13), pages 4773 - 4774, XP008175900
KURUMA Y ET AL.: "Question 7: biosynthesis of phosphatidic acid in liposome compartments - toward the self-reproduction of minimal cells", ORIG. LIFE EVOL. BIOSPH., vol. 37, no. 4-5, 2007, pages 409 - 413, XP019532459
NOIREAUX V ET AL.: "A vesicle bioreactor as a step toward an artificial cell assembly", PNAS, vol. 101, no. 51, 2004, pages 17669 - 17674, XP003003270
TAKEHIRO NISHIKAWA ET AL.: "Liposome o Hannoba to suru Idenshi Screening ni yoru Kasseigata Glucuronidase no Tansaku", ABSTRACTS, ANNUAL MEETING OF THE SOCIETY OF POLYMER SCIENCE, vol. 61, no. 1, 15 May 2012 (2012-05-15), pages ROMBUNNO.2G26, XP008175853
NISHIKAWA T ET AL.: "Construction of a gene screening system using giant unilamellar liposomes and a fluorescence-activated cell sorter", ANAL. CHEM., vol. 84, no. 11, 5 June 2012 (2012-06-05), pages 5017 - 5024, XP055181679
HARUKA SOGA ET AL.: "Construction of an in vitro gene screening system for membrane proteins", ABSTRACTS OF THE ANNUAL MEETING OF THE SOCIETY FOR BIOTECHNOLOGY, vol. 64, 25 September 2012 (2012-09-25), pages 197, XP008176026
NISHIKAWA T ET AL.: "Directed Evolution of Proteins through In Vitro Protein Synthesis in Liposomes", J. NUCLEIC ACIDS, vol. 2012, 2012, XP055181685
HOVIJITRA NT ET AL.: "Cell-free synthesis of functional aquaporin Z in synthetic liposomes", BIOTECHNOL. BIOENG., vol. 104, no. 1, 2009, pages 40 - 49, XP055069341
OHTSUKA T ET AL.: "Synthesis and in situ insertion of a site-specific fluorescently labeled membrane protein into cell-sized liposomes", ANAL. BIOCHEM., vol. 418, no. 1, 2011, pages 97 - 101, XP028277287
SUNAMI, T.; SATO, K.; MATSUURA, T.; TSUKADA, K.; URABE, I.; YOMO, T, ANALYTICAL BIOCHEMISTRY, vol. 357, 2006, pages 128 - 136
P. MUELLER; T. F. CHIEN, BIOPHYS. J., vol. 44, 1983, pages 375 - 381
MIGLENA I. ANGELOVE; DIMITER S. DIMITROV: "Faraday Discuss", CHEM. SOC., vol. 81, 1986, pages 303 - 311
See also references of EP 2876159A4
KURUMA Y ET AL.: "Question 7: biosynthesis of phosphatidic acid in liposome compartments - toward the self-reproduction of minimal cells", ORIG. LIFE EVOL. BIOSPH., vol. 37, no. 4-5, 2007, pages 409 - 413, XP019532459
NOIREAUX V ET AL.: "A vesicle bioreactor as a step toward an artificial cell assembly", PNAS, vol. 101, no. 51, 2004, pages 17669 - 17674, XP003003270
TAKEHIRO NISHIKAWA ET AL.: "Liposome o Hannoba to suru Idenshi Screening ni yoru Kasseigata Glucuronidase no Tansaku", ABSTRACTS, ANNUAL MEETING OF THE SOCIETY OF POLYMER SCIENCE, vol. 61, no. 1, 15 May 2012 (2012-05-15), pages ROMBUNNO.2G26, XP008175853
NISHIKAWA T ET AL.: "Construction of a gene screening system using giant unilamellar liposomes and a fluorescence-activated cell sorter", ANAL. CHEM., vol. 84, no. 11, 5 June 2012 (2012-06-05), pages 5017 - 5024, XP055181679
HARUKA SOGA ET AL.: "Construction of an in vitro gene screening system for membrane proteins", ABSTRACTS OF THE ANNUAL MEETING OF THE SOCIETY FOR BIOTECHNOLOGY, vol. 64, 25 September 2012 (2012-09-25), pages 197, XP008176026
NISHIKAWA T ET AL.: "Directed Evolution of Proteins through In Vitro Protein Synthesis in Liposomes", J. NUCLEIC ACIDS, vol. 2012, 2012, XP055181685
HOVIJITRA NT ET AL.: "Cell-free synthesis of functional aquaporin Z in synthetic liposomes", BIOTECHNOL. BIOENG., vol. 104, no. 1, 2009, pages 40 - 49, XP055069341
OHTSUKA T ET AL.: "Synthesis and in situ insertion of a site-specific fluorescently labeled membrane protein into cell-sized liposomes", ANAL. BIOCHEM., vol. 418, no. 1, 2011, pages 97 - 101, XP028277287
SUNAMI, T.; SATO, K.; MATSUURA, T.; TSUKADA, K.; URABE, I.; YOMO, T, ANALYTICAL BIOCHEMISTRY, vol. 357, 2006, pages 128 - 136
P. MUELLER; T. F. CHIEN, BIOPHYS. J., vol. 44, 1983, pages 375 - 381
MIGLENA I. ANGELOVE; DIMITER S. DIMITROV: "Faraday Discuss", CHEM. SOC., vol. 81, 1986, pages 303 - 311
See also references of EP 2876159A4
Attorney, Agent or Firm:
YAMAMOTO, Shusaku et al. (JP)
Shusaku Yamamoto (JP)
Shusaku Yamamoto (JP)
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