THE F1F0-ATPASE COMPLEX FROM BOVINE HEART-MITOCHONDRIA - THE MOLAR RATIO OF THE SUBUNITS IN THE STALK REGION LINKING THE F1 AND F-0 DOMAINS

The F-1 globular catalytic domain and the F-o intrinsic membrane domai n of the F1Fo-ATPases in bacteria, chloroplasts, and mitochondria are connected by a slender stalk. In the F1Fo complex from bovine heart mi tochondria, the stalk is thought to contain subunits OSCP, d, and F-6, and the globular part of the membrane bound subunit b, referred to as b'. It has been shown previously that the OSCP, b', d, and F-6 protei ns can be assembled in vitro into a water soluble complex named the '' stalk''. The stalk and F-1-ATPase together form another complex named F-1 . stalk. In this paper, the molar ratios of the OSCP, b (or b'), d , and F-6 in the stalk, F-1 . stalk, and F1Fo-ATPase complexes have be en investigated by three independent methods. By quantitation of radio activity incorporated by S-carboxymethylation with iodo-2-[(14)]acetic acid into a stalk complex containing a form of F-6 With the mutation Glu(3)-Cys, it was shown that the stalk consists of equimolar quantiti es of its four constituent proteins. In the stalk complex containing t he natural F-6 sequence, this conclusion was confirmed both by quantit ation of radioactivity incorporated by N-epsilon-acetimidation with et hyl [1-C-14]acetimidate, and by quantitative N-terminal sequence analy sis of subunits. By similar N-epsilon-acetimidation experiments, it ha s been demonstrated that the F-1 . stalk complex contains one copy per assembly of the OSCP, b', d, and F-6 proteins and that the F1Fo-ATPas e contains one copy per enzyme complex of subunits OSCP, b, and d. The presence of one copy per complex of the OSCP, b' (or b), d, and F-6 p roteins in the F-1 . stalk and F1Fo-ATPase complexes, respectively, wa s confirmed by quantitative sequencing.