Oxidative stress protein Oxr1 promotes V-ATPase holoenzyme disassembly in catalytic activity-independent manner

EMBO J. 2022 Feb 1;41(3):e109360. doi: 10.15252/embj.2021109360. Epub 2021 Dec 17.


The vacuolar ATPase (V-ATPase) is a rotary motor proton pump that is regulated by an assembly equilibrium between active holoenzyme and autoinhibited V1 -ATPase and Vo proton channel subcomplexes. Here, we report cryo-EM structures of yeast V-ATPase assembled in vitro from lipid nanodisc reconstituted Vo and mutant V1 . Our analysis identified holoenzymes in three active rotary states, indicating that binding of V1 to Vo provides sufficient free energy to overcome Vo autoinhibition. Moreover, the structures suggest that the unequal spacing of Vo 's proton-carrying glutamic acid residues serves to alleviate the symmetry mismatch between V1 and Vo motors, a notion that is supported by mutagenesis experiments. We also uncover a structure of free V1 bound to Oxr1, a conserved but poorly characterized factor involved in the oxidative stress response. Biochemical experiments show that Oxr1 inhibits V1 -ATPase and causes disassembly of the holoenzyme, suggesting that Oxr1 plays a direct role in V-ATPase regulation.

Keywords: Cryo-electron microscopy; Oxr1p; TLDc domain; reversible disassembly; vacuolar ATPase.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Cryoelectron Microscopy
  • Holoenzymes / chemistry
  • Mitochondrial Proteins / chemistry*
  • Mutagenesis
  • Oxidative Stress*
  • Protein Binding
  • Protein Multimerization
  • Saccharomyces cerevisiae Proteins / chemistry*
  • Saccharomyces cerevisiae Proteins / genetics
  • Vacuolar Proton-Translocating ATPases / chemistry
  • Vacuolar Proton-Translocating ATPases / genetics
  • Vacuolar Proton-Translocating ATPases / metabolism*


  • Holoenzymes
  • Mitochondrial Proteins
  • Oxr1 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Vph1 protein, S cerevisiae
  • Vacuolar Proton-Translocating ATPases

Associated data

  • PDB/7FDA
  • PDB/7FDB
  • PDB/7FDC
  • PDB/7FDE