Identification of target substrates of uncharacterized F-box proteins.

2014 Fiscal Year Final Research Report

• Project/Area Number: 24580152
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Applied biochemistry
• Research Institution: Tokyo Metropolitan Institute of Medical Science
• Principal Investigator: YOSHIDA Yukiko 公益財団法人東京都医学総合研究所, 生体分子先端研究分野, 主任研究員 (90271543)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Keywords: ユビキチン / ユビキチンリガーゼ / F-boxタンパク質 / プロテオミクス

Outline of Final Research Achievements

The Skp1-Cul1-F-box protein (SCF) complex, one of the best-characterized ubiquitin ligases, is composed of three invariable components and a variable component F-ox protein that serves as the substrate recognition subunit. Among the over 70 F-box proteins found in humans, less than half have been characterized. To identify substrates for uncharacterized F-box proteins, I developed a comprehensive method for identification of ubiquitinated substrates using the trypsin-resistant tandem ubiquitin-binding entity (TR-TUBE). The coexpression of TR-TUBE with a F-box protein stabilizes the ubiquitinated substrates by masking the ubiquitin chains. Using a combination of two strategies for enriching ubiquitinated substrates, TR-TUBE and ubiquitin Lys-e-Gly-Gly (diGly) remnant antibody, I successfully identified specific ubiquitin ligase-substrate pairs. Using this method, I identified target substrates of FBXO21, an uncharacterized and ubiquitiously expressed F-box protein.

Free Research Field

分子生物学・糖鎖生物学