Atpase Complex: Purification and Topology

Joshi, Saroj

Abstract

Present work suggests that mitochondrial Fo-F1, purified as described, has the highest reported Pi-ATP exchange activity. It is essentially free of adenylate kinase and phosphate transport protein. In highly resolving SDS-PAGE, it shows eighteen bands. The project is designed to purify it further to the minimum number of components and study the subunit arrangement and spatial relationship of various components. This would be achieved by using chemical as well as photo-affinity cross-linking reagents and the adducts will be identified with the help of site specific labels, subunit specific antibodies, and 2-dimensional SDS-PAGE, wherever applicable. It is hoped that the data would provide clues to the elucidation of mechanism of energy coupling processes that require intimate association of F1 and membrane components.

Funding Agency

• Agency: National Institute of Health (NIH)
• Institute: National Institute of General Medical Sciences (NIGMS)
• Type: Research Project (R01)
• Project #: 5R01GM026420-08
• Application #: 3273900
• Study Section: Physical Biochemistry Study Section (PB)
• Project Start: 1981-04-01
• Project End: 1991-03-31
• Budget Start: 1989-04-01
• Budget End: 1991-03-31
• Support Year: 8
• Fiscal Year: 1989

Institution

• Name: Boston Biomedical Research Institute
• DUNS #: 058893371
• City: Watertown
• State: MA
• Country: United States
• Zip Code: 02472

Publications

Joshi, S; Cao, G J; Nath, C et al. (1997) Oligomycin sensitivity conferring protein (OSCP) of bovine heart mitochondrial ATP synthase: high-affinity OSCP-Fo interactions require a local alpha-helix at the C-terminal end of the subunit. Biochemistry 36:10936-43

Joshi, S; Cao, G J; Nath, C et al. (1996) Oligomycin sensitivity conferring protein of mitochondrial ATP synthase: deletions in the N-terminal end cause defects in interactions with F1, while deletions in the C-terminal end cause defects in interactions with F0. Biochemistry 35:12094-103

Joshi, S; Javed, A A; Gibbs, L C (1992) Oligomycin sensitivity-conferring protein (OSCP) of mitochondrial ATP synthase. The carboxyl-terminal region of OSCP is essential for the reconstitution of oligomycin-sensitive H(+)-ATPase. J Biol Chem 267:12860-7

Joshi, S; Huang, Y G (1991) ATP synthase complex from bovine heart mitochondria: the oligomycin sensitivity conferring protein is essential for dicyclohexyl carbodiimide-sensitive ATPase. Biochim Biophys Acta 1067:255-8

Javed, A A; Joshi, S (1990) Targeted DNA sequencing: rapid identification of DNA clones by sequencing DNA using mixed oligodeoxynucleotide probes as primers. Biotechniques 9:28-32

Joshi, S; Burrows, R (1990) ATP synthase complex from bovine heart mitochondria. Subunit arrangement as revealed by nearest neighbor analysis and susceptibility to trypsin. J Biol Chem 265:14518-25

Pringle, M J; Kenneally, M K; Joshi, S (1990) ATP synthase complex from bovine heart mitochondria. Passive H+ conduction through F0 does not require oligomycin sensitivity-conferring protein. J Biol Chem 265:7632-7

Joshi, S; Pringle, M J (1989) ATP synthase complex from bovine heart mitochondria. Passive H+ conduction through mitochondrial coupling factor 6-depleted F0 complexes. J Biol Chem 264:15548-51

Joshi, S; Pringle, M J; Siber, R (1986) Topology and function of ""stalk"" proteins in the bovine mitochondrial H+-ATPase. J Biol Chem 261:10653-8

Joshi, S; Hughes, J B; Torok, K et al. (1985) Resolution and reconstitution of H+ -ATPase complex from beef heart mitochondria. Membr Biochem 5:309-25