title: Energy-driven subunit rotation at the interface between subunit a and the c oligomer in the F(O) sector of Escherichia coli ATP synthase.
abstract: Subunit rotation within the F(1) catalytic sector of the ATP synthase has been well documented, identifying the synthase as the smallest known rotary motor. In the membrane-embedded F(O) sector, it is thought that proton transport occurs at a rotor/stator interface between the oligomeric ring of c subunits (rotor) and the single-copy a subunit (stator). Here we report evidence for an energy-dependent rotation at this interface. F(O)F(1) was expressed with a pair of substituted cysteines positioned to allow an intersubunit disulfide crosslink between subunit a and a c subunit [aN214C/cM65C; Jiang, W. & Fillingame, R. H. (1998) Proc. Natl. Acad. Sci. USA 95, 6607--6612]. Membranes were treated with N,N'-dicyclohexyl-[(14)C]carbodiimide to radiolabel the D61 residue on less than 20% of the c subunits. After oxidation to form an a--c crosslink, the c subunit properly aligned to crosslink to subunit a was found to contain very little (14)C label relative to other members of the c ring. However, exposure to MgATP before oxidation significantly increased the radiolabel in the a-c crosslink, indicating that a different c subunit was now aligned with subunit a. This increase was not induced by exposure to MgADP/P(i). Furthermore, preincubation with MgADP and azide to inhibit F(1) or with high concentrations of N,N'-dicyclohexylcarbodiimide to label most c subunits prevented the ATP effect. These results provide evidence for an energy-dependent rotation of the c ring relative to subunit a.
authors: Department of Biochemistry and Molecular Biology, State University of New York Upstate Medical University, Syracuse, NY 13210, USA.
authors: M L
authors: ML
authors: Hutcheon
authors: T M
authors: TM
authors: Duncan
authors: H
authors: H
authors: Ngai
authors: R L
authors: RL
authors: Cross
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D003432:Cross-Linking Reagents; D004024:Dicyclohexylcarbodiimide / Q000378:metabolism; D004735:Energy Transfer; D004791:Enzyme Inhibitors / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D011487:Protein Conformation; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism
chemical_list: D003432:Cross-Linking Reagents; D004791:Enzyme Inhibitors; D004024:Dicyclohexylcarbodiimide; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases