Coordinated conformational changes in the V1 complex during V-ATPase reversible dissociation

Vacuolar-type ATPases (V-ATPases) are rotary enzymes that acidify intracellular compartments in eukaryotic cells. These multi-subunit complexes consist of a cytoplasmic V 1 region that hydrolyzes ATP and a membrane-embedded V O region that transports protons. V-ATPase activity is regulated by reversible dissociation of the two regions, with the isolated V 1 and V O complexes becoming autoinhibited on disassembly and subunit C subsequently detaching from V 1 . In yeast, assembly of the V 1 and V O regions is mediated by the regulator of the ATPase of vacuoles and endosomes (RAVE) complex through an unknown mechanism. We used cryogenic-electron microscopy of yeast V-ATPase to determine structures of the intact enzyme, the dissociated but complete V 1 complex and the V 1 complex lacking subunit C. On separation, V 1 undergoes a dramatic conformational rearrangement, with its rotational state becoming incompatible for reassembly with V O . Loss of subunit C allows V 1 to match the rotational state of V O , suggesting how RAVE could reassemble V 1 and V O by recruiting subunit C. V-ATPases acidify the intracellular compartments of eukaryotic cells and their activity is regulated by reversible dissociation of the complex. Cryo-EM structures show the conformational changes associated with assembly and autoinhibition of V-ATPase.