In the ubiquitin (Ub)/proteasome proteolytic pathway, substrate-linked polyUb is the specific targeting signal that leads proteins to be degraded by the multisubunit ATP-dependent protease complex called the 26S proteasome. The mechanisms by which the proteasome recognizes, unfolds, and translocates the substrate into its internal proteolytic chamber are poorly understood. In the proposal, in vitro biochemistry augmented by yeast genetics will be used to address how key events such as substrate unfolding and translocation are orchestrated by the six different ATPase subunits of the 19S regulatory complex of 26S proteasomes. For this purpose, well-defined polyUb-conjugates of wild-type and destabilized variants of two model proteins, dihydrofolate reductase and titin I27 domain, have been developed. By using these substrates to measure degradation and ATPase activities with purified wild-type and ATPase-mutated 26S proteasomes, we will be able to resolve the contributions that ATP hydrolysis and, potentially, individual ATPase subunits make toward the substrate binding, unfolding, and translocation steps of the degradation reaction. Our results will provide a basic understanding of how the 26S proteasome complex uses ATP to promote proteolysis. Overall, the results of the proposed research will significantly enhance our understanding of how the 26S proteasome degrades proteins and how the complex is regulated. The significance of proteasome research to human health has been highlighted by the recent FDA approval of the use of proteasome inhibitor as a treatment for multiple myeloma.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM073028-03
Application #
7576697
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Ikeda, Richard A
Project Start
2007-05-01
Project End
2012-02-28
Budget Start
2009-03-01
Budget End
2010-02-28
Support Year
3
Fiscal Year
2009
Total Cost
$312,093
Indirect Cost
Name
Weill Medical College of Cornell University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
060217502
City
New York
State
NY
Country
United States
Zip Code
10065