Title:
METHOD OF SCREENING TARGET PROTEIN OF UBIQUITINATION BY UBIQUITIN LIGASE
Document Type and Number:
WIPO Patent Application WO/2006/035975
Kind Code:
A1
Abstract:
It is intended to develop an experiment system for screening a protein to be ubiquitinated by ubiquitin ligase and identify a target protein of the ubiquitination by using the system. It is found out that a target protein can be stabilized and consequently screened by, in a yeast two-hybrid screening system, (1) using, as a bait, not the full-length F-box protein but a mutated F-box protein lacking the F-box motif, which is seemingly an Skp1-binding region, either as a whole or to such an extent as losing the ability to bind to Skp1, and (2) further using a mutant having a lowered ability to degrade the target protein (for example, a temperature-sensitive mutant or a deletion mutant).
Inventors:
KISHI TSUTOMU (JP)
Application Number:
PCT/JP2005/018245
Publication Date:
April 06, 2006
Filing Date:
September 27, 2005
Export Citation:
Assignee:
RIKEN (JP)
KISHI TSUTOMU (JP)
KISHI TSUTOMU (JP)
International Classes:
C12Q1/68; C12N15/09; G01N33/15; G01N33/50; G01N33/68
Other References:
TOKARZ S ET AL: "The ISG15 isopeptidase UBP43 is regulated by proteolysis via the SCFSkp2 ubiquitin ligase.", J BIOL CHEM., vol. 279, no. 45, 5 November 2004 (2004-11-05), pages 46424 - 46430, XP002998910
LI Z ET AL: "Ubiquitination of a novel deubiquitinating enzyme requires direct binding to von Hippel-Lindau tumor suppressor protein.", J BIOL CHEM., vol. 277, no. 7, 15 February 2002 (2002-02-15), pages 4656 - 4662, XP002296282
KAMURA T ET AL: "A molecular basis for stabilization of the von Hippel-Lindau (VHL) tumor suppressor protein by components of the VHL ubiquitin ligase.", J BIOL CHEM., vol. 277, no. 33, 16 August 2002 (2002-08-16), pages 30388 - 30393, XP002998911
LI Y ET AL: "Stability of homologue of Slimb F-box protein is regulated by availability of its substrate.", J BIOL CHEM., vol. 279, no. 12, 19 March 2004 (2004-03-19), pages 11074 - 11080, XP002998912
COCKMAN M E ET AL: "Hypoxia inducible factor-alpha binding and ubiquitylation by the von Hippel-Lindau tumor suppressor protein.", J BIOL CHEM., vol. 275, no. 33, 18 August 2000 (2000-08-18), pages 25733 - 25741, XP002998913
BAI C ET AL: "SKP1 connects cell cycle regulators to the ubiquitin proteolysis machinery through a novel motif, the F-box.", CELL., vol. 86, no. 2, 26 July 1996 (1996-07-26), pages 263 - 274, XP002082118
SCHWAB M ET AL: "Yeast Hct1 recognizes the mitotic cyclin Clb2 and other substrates of the ubiquitin ligase APC.", EMBO J., vol. 20, no. 18, 17 September 2001 (2001-09-17), pages 5165 - 5175, XP002998914
KISHI TSUTOMU ET AL: "SCF Ubiquitin Ligase no Hyoteki Tanpakushitsu o Dotei suru Tameno Jikken Shuho no Kaihatsu.", 27TH ANNUAL MEETING OF THE MOLECULAR BIOLOGY SOCIETY OF JAPAN PROGRAM KOEN YOSHISHU., 25 November 2004 (2004-11-25), pages 546, 1PB-173, XP002998915
LI Z ET AL: "Ubiquitination of a novel deubiquitinating enzyme requires direct binding to von Hippel-Lindau tumor suppressor protein.", J BIOL CHEM., vol. 277, no. 7, 15 February 2002 (2002-02-15), pages 4656 - 4662, XP002296282
KAMURA T ET AL: "A molecular basis for stabilization of the von Hippel-Lindau (VHL) tumor suppressor protein by components of the VHL ubiquitin ligase.", J BIOL CHEM., vol. 277, no. 33, 16 August 2002 (2002-08-16), pages 30388 - 30393, XP002998911
LI Y ET AL: "Stability of homologue of Slimb F-box protein is regulated by availability of its substrate.", J BIOL CHEM., vol. 279, no. 12, 19 March 2004 (2004-03-19), pages 11074 - 11080, XP002998912
COCKMAN M E ET AL: "Hypoxia inducible factor-alpha binding and ubiquitylation by the von Hippel-Lindau tumor suppressor protein.", J BIOL CHEM., vol. 275, no. 33, 18 August 2000 (2000-08-18), pages 25733 - 25741, XP002998913
BAI C ET AL: "SKP1 connects cell cycle regulators to the ubiquitin proteolysis machinery through a novel motif, the F-box.", CELL., vol. 86, no. 2, 26 July 1996 (1996-07-26), pages 263 - 274, XP002082118
SCHWAB M ET AL: "Yeast Hct1 recognizes the mitotic cyclin Clb2 and other substrates of the ubiquitin ligase APC.", EMBO J., vol. 20, no. 18, 17 September 2001 (2001-09-17), pages 5165 - 5175, XP002998914
KISHI TSUTOMU ET AL: "SCF Ubiquitin Ligase no Hyoteki Tanpakushitsu o Dotei suru Tameno Jikken Shuho no Kaihatsu.", 27TH ANNUAL MEETING OF THE MOLECULAR BIOLOGY SOCIETY OF JAPAN PROGRAM KOEN YOSHISHU., 25 November 2004 (2004-11-25), pages 546, 1PB-173, XP002998915
Attorney, Agent or Firm:
Hiraki, Yusuke (3-20 Toranomon 4-chom, Minato-ku Tokyo, JP)
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