Defining the domain of binding of F1 subunit epsilon with the polar loop of F0 subunit c in the Escherichia coli ATP synthase.
We have previously shown that the E31C-substituted epsilon subunit of F1 can be cross-linked by disulfide bond formation to the Q42C-substituted c subunit of F0 in the Escherichia coli F1F0-ATP synthase complex (Zhang, Y., and Fillingame, R. H. (1995) J. Biol. Chem. 270, 24609-24614). The interactions of subunits epsilon and c are thought to be central to the coupling of H+ transport through F0 to ATP synthesis in F1. To further define the domains of interaction, we have introduced additional Cys into subunit epsilon and subunit c and tested for cross-link formation following sulfhydryl oxidation. The results show that Cys, in a continuous stretch of residues 26-33 in subunit epsilon, can be cross-linked to Cys at positions 40, 42, and 44 in the polar loop region of subunit c. The results are interpreted, and the subunit interaction is modeled using the NMR and x-ray diffraction structures of the monomeric subunits together with information on the packing arrangement of subunit c in a ring of 12 subunits. In the model, residues 26-33 form a turn of antiparallel beta-sheet which packs between the polar loop regions of adjacent subunit c at the cytoplasmic surface of the c12 oligomer.
Hermolin
J
J
Department of Biomolecular Chemistry, University of Wisconsin Medical School, Madison, Wisconsin 53706, USA.
Dmitriev
O Y
OY
Zhang
Y
Y
Fillingame
R H
RH
D001665:Binding Sites; D003432:Cross-Linking Reagents; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D004220:Disulfides; D004926:Escherichia coli / Q000201:enzymology*; D025261:Mitochondrial Proton-Translocating ATPases; D008958:Models, Molecular; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000097795:ATPase Inhibitory Protein
D003432:Cross-Linking Reagents; D003545:Cysteine; D004220:Disulfides; D025261:Mitochondrial Proton-Translocating ATPases; D011506:Proteins; D006180:Proton-Translocating ATPases; D000097795:ATPase Inhibitory Protein; C103607:mitochondrial ATPase subunit c