title: Membrane embedded location of Na+ or H+ binding sites on the rotor ring of F1F0 ATP synthases.
abstract: Recent crosslinking studies indicated the localization of the coupling ion binding site in the Na+-translocating F1F0 ATP synthase of Ilyobacter tartaricus within the hydrophobic part of the bilayer. Similarly, a membrane embedded H+-binding site is accepted for the H+-translocating F1F0 ATP synthase of Escherichia coli. For a more definite analysis, we performed parallax analysis of fluorescence quenching with ATP synthases from both I. tartaricus and E. coli. Both ATP synthases were specifically labelled at their c subunit sites with N-cyclohexyl-N'-(1-pyrenyl)carbodiimide, a fluorescent analogue of dicyclohexylcarbodiimide and the enzymes were reconstituted into proteoliposomes. Using either soluble quenchers or spinlabelled phospholipids, we observed a deeply membrane embedded binding site, which was quantitatively determined for I. tartaricus and E. coli to be 1.3 +/- 2.4 A and 1.8 +/- 2.8 A from the bilayer center apart, respectively. These data show a conserved topology among enzymes of different species. We further demonstrated the direct accessibility for Na+ ions to the binding sites in the reconstituted I. tartaricus c11 oligomer in the absence of any other subunits, pointing to intrinsic rotor channels. The common membrane embedded location of the binding site of ATP synthases suggest a common mechanism for ion transfer across the membrane.
authors.affiliation: Institut für Mikrobiologie der Eidgenössischen Technischen Hochschule, ETH Zentrum, Zürich, Switzerland.
authors.forename: Christoph
authors.initials: C
authors.lastname: von Ballmoos
authors.forename: Thomas
authors.initials: T
authors.lastname: Meier
authors.forename: Peter
authors.initials: P
authors.lastname: Dimroth
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D025243:Bacterial Proton-Translocating ATPases / Q000737:chemistry*; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005673:Fusobacterium / Q000378:metabolism; D006859:Hydrogen / Q000737:chemistry*; D006868:Hydrolysis; D008954:Models, Biological; D011485:Protein Binding; D012964:Sodium / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors
chemical_list: D006859:Hydrogen; D000255:Adenosine Triphosphate; D012964:Sodium; D025243:Bacterial Proton-Translocating ATPases