title: Atomic model for the membrane-embedded VOmotor of a eukaryotic V-ATPase.
abstract: Vacuolar-type ATPases (V-ATPases) are ATP-powered proton pumps involved in processes such as endocytosis, lysosomal degradation, secondary transport, TOR signalling, and osteoclast and kidney function. ATP hydrolysis in the soluble catalytic V1region drives proton translocation through the membrane-embedded VOregion via rotation of a rotor subcomplex. Variability in the structure of the intact enzyme has prevented construction of an atomic model for the membrane-embedded motor of any rotary ATPase. We induced dissociation and auto-inhibition of the V1and VOregions of the V-ATPase by starving the yeast Saccharomyces cerevisiae, allowing us to obtain a ~3.9-Å resolution electron cryomicroscopy map of the VOcomplex and build atomic models for the majority of its subunits. The analysis reveals the structures of subunits ac8c'c″de and a protein that we identify and propose to be a new subunit (subunit f). A large cavity between subunit a and the c-ring creates a cytoplasmic half-channel for protons. The c-ring has an asymmetric distribution of proton-carrying Glu residues, with the Glu residue of subunit c″ interacting with Arg735 of subunit a. The structure suggests sequential protonation and deprotonation of the c-ring, with ATP-hydrolysis-driven rotation causing protonation of a Glu residue at the cytoplasmic half-channel and subsequent deprotonation of a Glu residue at a luminal half-channel.
authors: Molecular Structure and Function Program, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada.
authors: Mohammad T
authors: MT
authors: Mazhab-Jafari
authors: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia 20147, USA.
authors: Alexis
authors: A
authors: Rohou
authors: Department of Chemistry, Physical and Theoretical Chemistry Laboratory, University of Oxford, Oxford OX1 3QZ, UK.
authors: Carla
authors: C
authors: Schmidt
authors: Molecular Structure and Function Program, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada.
authors: Stephanie A
authors: SA
authors: Bueler
authors: Molecular Structure and Function Program, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada.
authors: Samir
authors: S
authors: Benlekbir
authors: Department of Chemistry, Physical and Theoretical Chemistry Laboratory, University of Oxford, Oxford OX1 3QZ, UK.
authors: Carol V
authors: CV
authors: Robinson
authors: Molecular Structure and Function Program, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada.|Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 1L7, Canada.|Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
authors: John L
authors: JL
authors: Rubinstein
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D001120:Arginine / Q000737:chemistry / Q000378:metabolism; D020285:Cryoelectron Microscopy; D018698:Glutamic Acid / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D008958:Models, Molecular; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011522:Protons; D012399:Rotation; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D025262:Vacuolar Proton-Translocating ATPases / Q000737:chemistry* / Q000648:ultrastructure*
chemical_list: D021122:Protein Subunits; D011522:Protons; D029701:Saccharomyces cerevisiae Proteins; D018698:Glutamic Acid; D000255:Adenosine Triphosphate; D001120:Arginine; D025262:Vacuolar Proton-Translocating ATPases