tailieunhanh - Báo cáo khoa học: Dynamic interactions of proteins in complex networks: identifying the complete set of interacting E2s for functional investigation of E3-dependent protein ubiquitination

A ubiquitin ligase (E3) functions at the crossroad between ubiquitin acti-vation and the attachment of ubiquitin to protein substrates. During this process, the E3 interacts with both a substrate and a ubiquitin-conjugat-ing enzyme (E2). Although a major goal when investigating an E3 is to identify its substrates, recent evidence indicates that the E2 dictates the type of ubiquitin modification that will occur on the substrate. | ỊFEBS Journal MINIREVIEW Dynamic interactions of proteins in complex networks identifying the complete set of interacting E2s for functional investigation of E3-dependent protein ubiquitination Devin E. Christensen1 and Rachel E. Klevit2 1 Department of Biochemistry University of Utah Salt Lake City UT USA 2 Department of Biochemistry University of Washington Seattle WA USA Keywords BRCA1 NMR protein-protein interactions RING domain UbcH5 Ubc13 ubiquitin ligase ubiquitination ubiquitin-conjugating enzyme yeast two-hybrid Correspondence R. E. Klevit Department of Biochemistry University of Washington Box 357350 1705 NE Pacific Street Seattle WA 98195 USA Fax 1 206 543 8394 Tel 1 206 543 5891 E-mail klevit@ Received 29 May 2009 revised 8 July 2009 accepted 15 July 2009 doi A ubiquitin ligase E3 functions at the crossroad between ubiquitin activation and the attachment of ubiquitin to protein substrates. During this process the E3 interacts with both a substrate and a ubiquitin-conjugat-ing enzyme E2 . Although a major goal when investigating an E3 is to identify its substrates recent evidence indicates that the E2 dictates the type of ubiquitin modification that will occur on the substrate. There are 30 E2s identified in the human genome many of which remain to be characterized. We found that the RING E3 BRCA1 BARD1 can interact with 10 different E2s. The ability of BRCA1 to interact with multiple E2s is likely to be a common feature among other RING and U-box E3s. We and others have also found that certain E2s show a preference for attaching either the first ubiquitin to a substrate lysine or ubiquitin to itself chain building suggesting that E2s may play a role in dictating product formation. Therefore when investigating the functions of an E3 it is advisable to identify all E2s that interact with the E3 so that these can be used in E3-dependent substrate-ubiquitination assays. We describe a method used to identify .