Interactions between the oligomycin sensitivity conferring protein (OSCP) and beef heart mitochondrial F1-ATPase. 2. Identification of the interacting F1 subunits by cross-linking.
Interactions between oligomycin sensitivity conferring protein (OSCP) and subunits of beef heart mitochondrial F1-ATPase have been explored by cross-linking at an OSCP/F1 molar ratio close to 1 to ensure specific high-affinity binding of OSCP to F1 [see Dupuis et al. [Dupuis, A., Issartel, J.-P., Lunardi, J., Satre, M., & Vignais, P.V. (1985) Biochemistry (preceding paper in this issue)]]. Cross-links between F1 subunits and OSCP were established by means of two zero length cross-linkers, 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide and N-(ethoxycarbonyl)-2-ethoxydihydroquinoline. The cross-linked products were separated by sodium dodecyl suflate-polyacrylamide gel electrophoresis. Coomassie blue staining revealed two cross-linked products of Mr 75 000 and 80 000 which could result from the binding of OSCP to the alpha and beta subunits of F1. Definite identification of the cross-linked products was achieved by chemical labeling with specific radiolabeled reagents and by blotting on nitrocellulose filters followed by immunocharacterization with anti-alpha, anti-beta, and anti-OSCP antibodies. OSCP was found to cross-link with the alpha and beta subunits of F1.
Dupuis
A
A
Lunardi
J
J
Issartel
J P
JP
Vignais
P V
PV
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D003432:Cross-Linking Reagents; D005022:Ethyldimethylaminopropyl Carbodiimide; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000378:metabolism*; D025261:Mitochondrial Proton-Translocating ATPases; D008970:Molecular Weight; D009840:Oligomycins / Q000378:metabolism*; D011485:Protein Binding; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D011804:Quinolines
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D008565:Membrane Proteins; D009840:Oligomycins; D011804:Quinolines; C000470:EEDQ; D000251:Adenosine Triphosphatases; D025261:Mitochondrial Proton-Translocating ATPases; D006180:Proton-Translocating ATPases; C020439:oligomycin sensitivity-conferring protein; D005022:Ethyldimethylaminopropyl Carbodiimide