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TitleCryo-EM and MD infer water-mediated proton transport and autoinhibition mechanisms of V complex.
Journal, issue, pagesSci Adv, Vol. 6, Issue 41, Year 2020
Publish dateOct 7, 2020
AuthorsSoung-Hun Roh / Mrinal Shekhar / Grigore Pintilie / Christophe Chipot / Stephan Wilkens / Abhishek Singharoy / Wah Chiu /
PubMed AbstractRotary vacuolar adenosine triphosphatases (V-ATPases) drive transmembrane proton transport through a V proton channel subcomplex. Despite recent high-resolution structures of several rotary ATPases, ...Rotary vacuolar adenosine triphosphatases (V-ATPases) drive transmembrane proton transport through a V proton channel subcomplex. Despite recent high-resolution structures of several rotary ATPases, the dynamic mechanism of proton pumping remains elusive. Here, we determined a 2.7-Å cryo-electron microscopy (cryo-EM) structure of yeast V proton channel in nanodisc that reveals the location of ordered water molecules along the proton path, details of specific protein-lipid interactions, and the architecture of the membrane scaffold protein. Moreover, we uncover a state of V that shows the -ring rotated by ~14°. Molecular dynamics simulations demonstrate that the two rotary states are in thermal equilibrium and depict how the protonation state of essential glutamic acid residues couples water-mediated proton transfer with -ring rotation. Our cryo-EM models and simulations also rationalize a mechanism for inhibition of passive proton transport as observed for free V that is generated as a result of V-ATPase regulation by reversible disassembly in vivo.
External linksSci Adv / PubMed:33028525 / PubMed Central
MethodsEM (single particle)
Resolution2.7 - 3.6 Å
Structure data

30034

6m0r

EMDB-30034, PDB-6m0r:
2.7A Yeast Vo state3
Method: EM (single particle) / Resolution: 2.7 Å

30035

6m0s

EMDB-30035, PDB-6m0s:
3.6A Yeast Vo state3 prime
Method: EM (single particle) / Resolution: 3.6 Å

Chemicals

ChemComp-PEE:
1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / DOPE, phospholipid*YM / Discrete optimized protein energy

ChemComp-PPV:
PYROPHOSPHATE / Pyrophosphate

ChemComp-EYR:
(6~{E},10~{E},14~{E},18~{E},22~{E},26~{E},30~{R})-2,6,10,14,18,22,26,30-octamethyldotriaconta-2,6,10,14,18,22,26-heptaene

ChemComp-HOH:
WATER / Water

Source
  • Saccharomyces cerevisiae (brewer's yeast)
  • saccharomyces cerevisiae (strain atcc 204508 / s288c) (yeast)
KeywordsTRANSPORT PROTEIN / V-ATPase / Vo sub-complex / CryoEM / rotary motor

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