Structural snapshots of V/A-ATPase reveal the rotary catalytic mechanism of rotary ATPases.

V/A-ATPase is a motor protein that shares a common rotary catalytic mechanism with F o F 1 ATP synthase. When powered by ATP hydrolysis, the V 1 domain rotates the central rotor against the A 3 B 3 hexamer, composed of three catalytic AB dimers adopting different conformations (AB open , AB semi , and AB closed ). Here, we report the atomic models of 18 catalytic intermediates of the V 1 domain of V/A-ATPase under different reaction conditions, determined by single particle cryo-EM. The models reveal that the rotor does not rotate immediately after binding of ATP to the V 1 . Instead, three events proceed simultaneously with the 120˚ rotation of the shaft: hydrolysis of ATP in AB semi , zipper movement in AB open by the binding ATP, and unzipper movement in AB closed with release of both ADP and Pi. This indicates the unidirectional rotation of V/A-ATPase by a ratchet-like mechanism owing to ATP hydrolysis in AB semi , rather than the power stroke model proposed previously for F 1 -ATPase.