The focus of this proposal is on the mitochondrial ATP synthase (FoF1- ATPase), one of nature's most important, unique, and yet poorly understood molecular mechanisms. The immediate objectives are to understand both the mechanism by which this enzyme makes ATP at a chemical level and the role of the recently discovered """"""""motor"""""""" function of the enzyme in this process. Progress on this project has been substantial with 42 papers resulting during the MERIT award period (1988-1998) including 25 published in, or prepared for, refereed journals. Prominent among the most recent are those describing a) the formation of a transition state-like complex on the pathway for ATP synthesis; b) the atomic resolution structure of the F/1 moiety of rat liver ATP synthase; c) the properties of the delta and OSCP subunits, putative """"""""rotor"""""""" and """"""""stator"""""""" components, respectively, of the motor involved in ATP synthase function; and d) a novel method for rapidly screening detergents for their use in membrane protein crystallization trials. These recent studies set the stage for the design and vigorous pursuit of all experiments deemed essential to meet the objective of this proposal.
Specific Aims are four-fold and will be to: 1. Establish the extent to which conformational changes occur in the active site of the F/1 moiety as the ATP synthase reaction proceeds from the substrate bound states to the transition state, and define the role of the critical """"""""P-loop"""""""" alanine of the beta subunit. 2. Identify interactions of the """"""""structurally elusive"""""""" F1-delta subunit with other F1 subunits during the ATP synthase reaction, and develop a motility assay to determine whether this subunit physically rotates during catalysis. 3. Gain greater insight into the role of the OSCP subunit in ATP synthase function by better defining its interactions with the beta and other FoF1 subunits during catalysis, and determine whether OSCP and/or other subunits comprise the putative """"""""stator-like"""""""" structural elements recently observed in electron micrographs. 4. Extend, with determination, vigor, and passion, our recently successful collaborative efforts to obtain 3-dimensional structural information about the ATP synthase, from the F1 moiety to the complete complex. The studies proposed here are not only fundamental to our understanding of ATP synthase function, but will be critical to our future understanding of its dysfunction, e.g., in cancer and aging, where ATP synthesis is down-regulated, and a whole host of pathologies generally categorized as """"""""mitochondrial diseases"""""""".

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA010951-31A1
Application #
2769722
Study Section
Special Emphasis Panel (ZRG3-BBCB (02))
Program Officer
Gallahan, Daniel L
Project Start
1979-05-01
Project End
2003-11-30
Budget Start
1998-12-15
Budget End
1999-11-30
Support Year
31
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Johns Hopkins University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
045911138
City
Baltimore
State
MD
Country
United States
Zip Code
21218
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