title: The transmembrane domain of subunit b of the Escherichia coli F1F(O) ATP synthase is sufficient for H(+)-translocating activity together with subunits a and c.
abstract: Subunit b is indispensable for the formation of a functional H(+)-translocating F(O) complex both in vivo and in vitro. Whereas the very C-terminus of subunit b interacts with F(1) and plays a crucial role in enzyme assembly, the C-terminal region is also considered to be necessary for proper reconstitution of F(O) into liposomes. Here, we show that a synthetic peptide, residues 1-34 of subunit b (b(1-34)) [Dmitriev, O., Jones, P.C., Jiang, W. & Fillingame, R.H. (1999) J. Biol. Chem.274, 15598-15604], corresponding to the membrane domain of subunit b was sufficient in forming an active F(O) complex when coreconstituted with purified ac subcomplex. H(+) translocation was shown to be sensitive to the specific inhibitor N,N'-dicyclohexylcarbodiimide, and the resulting F(O) complexes were deficient in binding of isolated F(1). This demonstrates that only the membrane part of subunit b is sufficient, as well as necessary, for H(+) translocation across the membrane, whereas the binding of F(1) to F(O) is mainly triggered by C-terminal residues beyond Glu34 in subunit b. Comparison of the data with former reconstitution experiments additionally indicated that parts of the hydrophilic portion of the subunit b dimer are not involved in the process of ion translocation itself, but might organize subunits a and c in F(O) assembly. Furthermore, the data obtained functionally support the monomeric NMR structure of the synthetic b(1-34).
authors.affiliation: Universität Osnabrück, Fachbereich Biologie/Chemie, Abteilung Mikrobiologie, Osnabrück, Germany. joerg.greie@biologie.uni-osnabrueck.de
authors.forename: Jörg-Christian
authors.initials: JC
authors.lastname: Greie
authors.forename: Thomas
authors.initials: T
authors.lastname: Heitkamp
authors.forename: Karlheinz
authors.initials: K
authors.lastname: Altendorf
mesh_terms: D001692:Biological Transport; D004024:Dicyclohexylcarbodiimide / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism; D046911:Macromolecular Substances; D025261:Mitochondrial Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011522:Protons
chemical_list: D029968:Escherichia coli Proteins; D008081:Liposomes; D046911:Macromolecular Substances; D010455:Peptides; D021122:Protein Subunits; D011522:Protons; D004024:Dicyclohexylcarbodiimide; C474451:F1F0-ATP synthase; D025261:Mitochondrial Proton-Translocating ATPases