Membrane topography and near-neighbor relationships of the mitochondrial ATP synthase subunits e, f, and g.
The well characterized subunits of the bovine ATP synthase complex are the alpha, beta, gamma, delta, and epsilon subunits of the catalytic sector, F1; the ATPase inhibitor protein; and subunits a, b, c, and d, OSCP (oligomycin sensitivity-conferring protein), F6, and A6L, which are present in the membrane sector, F0, and the 45-A-long stalk that connects F1 to F0. It has been shown recently that bovine ATP synthase preparations also contain three small polypeptides, designated e, f, and g, with respective molecular masses of 8.2, 10. 2, and 11.3 kDa. To ascertain their involvement as bona fide subunits of the ATP synthase and to investigate their membrane topography and proximity to the above ATP synthase subunits, polyclonal antipeptide antibodies were raised in the rabbit to the COOH-terminal amino acid residues 57-70 of e, 75-86 of f, and 91-102 of g. It was shown that (i) e, f, and g could be immunoprecipitated with anti-OSCP IgG from a fraction of bovine submitochondrial particles enriched in oligomycin-sensitive ATPase; (ii) the NH2 termini of f and g are exposed on the matrix side of the mitochondrial inner membrane and can be curtailed by proteolysis; (iii) the COOH termini of all three polypeptides are exposed on the cytosolic side of the inner membrane; and (iv) f cross-links to A6L and to g, and e cross-links to g and appears to form an e-e dimer. Thus, the bovine ATP synthase complex appears to have 16 unlike subunits, twice as many as its counterpart in Escherichia coli.
Belogrudov
G I
GI
Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037, USA.
Tomich
J M
JM
Hatefi
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002918:Chymotrypsin / Q000378:metabolism; D003432:Cross-Linking Reagents; D007158:Immunologic Techniques; D046911:Macromolecular Substances; D008929:Mitochondria, Heart / Q000201:enzymology* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D008970:Molecular Weight; D006180:Proton-Translocating ATPases / Q000737:chemistry / Q000648:ultrastructure*; D012995:Solubility; D014357:Trypsin / Q000378:metabolism
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D002918:Chymotrypsin; D014357:Trypsin; D006180:Proton-Translocating ATPases