Vacuolar ATPases (V‐ATPases) are essential proton pumps that acidify the lumen of subcellular organelles in all eukaryotic cells and the extracellular space in some tissues. V‐ATPase activity is regulated by a unique mechanism referred to as reversible disassembly, wherein the soluble catalytic sector, V1, is released from the membrane and its MgATPase activity silenced. The crystal structure of yeast V1 presented here shows that activity silencing involves a large conformational change of subunit H, with its C‐terminal domain rotating ~150° from a position near the membrane in holo V‐ATPase to a position at the bottom of V1 near an open catalytic site. Together with biochemical data, the structure supports a mechanistic model wherein subunit H inhibits ATPase activity by stabilizing an open catalytic site that results in tight binding of inhibitory ADP at another site.
The proton‐pumping vacuolar ATPase is regulated by reversible disassembly of subcomplexes V1 and V0. The crystal structure of V1 reveals the basis for autoinhibition and provides a mechanism for V‐ATPase regulation.
The crystal structure of autoinhibited yeast V1‐ATPase was solved at 6.2–6.5 Å resolution.
Autoinhibition of V1‐ATPase activity involves a large‐scale domain rotation (150°) of the H subunit.
The conformational change in the H subunit appears to stabilize inhibitory ADP in one of the three catalytic sites.
The structural changes observed in V1 upon enzyme disassembly reveal the mechanism of V‐ATPase regulation.
- Received November 6, 2015.
- Revision received May 11, 2016.
- Accepted May 12, 2016.
- © 2016 The Authors
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