Yeast V-ATPase Proteolipid Ring Acts as a Large-conductance Transmembrane Protein Pore.
The vacuolar H(+)-ATPase (V-ATPase) is a rotary motor enzyme that acidifies intracellular organelles and the extracellular milieu in some tissues. Besides its canonical proton-pumping function, V-ATPase's membrane sector, Vo, has been implicated in non-canonical functions including membrane fusion and neurotransmitter release. Here, we report purification and biophysical characterization of yeast V-ATPase c subunit ring (c-ring) using electron microscopy and single-molecule electrophysiology. We find that yeast c-ring forms dimers mediated by the c subunits' cytoplasmic loops. Electrophysiology measurements of the c-ring reconstituted into a planar lipid bilayer revealed a large unitary conductance of ~8.3 nS. Thus, the data support a role of V-ATPase c-ring in membrane fusion and neuronal communication.
Department of Biochemistry &Molecular Biology, SUNY Upstate Medical University, Syracuse, New York 13210, USA.
Sergio
S
Couoh-Cardel
Department of Physics, Syracuse University, 201 Physics Bldg., Syracuse, New York 13244-1130, USA.
Yi-Ching
YC
Hsueh
Department of Biochemistry &Molecular Biology, SUNY Upstate Medical University, Syracuse, New York 13210, USA.
Stephan
S
Wilkens
Department of Physics, Syracuse University, 201 Physics Bldg., Syracuse, New York 13244-1130, USA.|Structural Biology, Biochemistry, and Biophysics Program, Syracuse University, 111 College Place, Syracuse, New York 13244-4100, USA.|The Syracuse Biomaterials Institute, Syracuse University, 318 Bowne Hall, Syracuse, New York 13244-1200, USA.
Liviu
L
Movileanu
D004553:Electric Conductivity; D055724:Electrophysiological Phenomena; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D054730:Protein Interaction Domains and Motifs; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D015003:Yeasts / Q000502:physiology*
D008565:Membrane Proteins; D021122:Protein Subunits; D025262:Vacuolar Proton-Translocating ATPases