Catalytic dwell oscillations complete the F1-ATPase mechanism.

Abstract

The F₁-ATPase molecular motor rotates subunit-γ in 120° power strokes within its ring of three catalytic sites separated by catalytic dwells for ATP hydrolysis and Pi release. By monitoring rotary position of subunit-γ in E. coli F₁ every 5 μs, we resolved Stage-1 catalytic dwell oscillations that extend from -13° to 13° centered at 0° consistent with F₁ structures containing transition state inhibitors, which decay by a first order process consistent with ATP hydrolysis. During Stage-2, 80% of the oscillations extend from 3° and 25° centered at 14°, while 20% are centered at 33° and can extend to 27°-44° comparable to the ATP binding position. Remarkably, in Stage-3 subunit-γ returns to 0° to end the catalytic dwell, which keeps the start of power strokes in phase for consecutive rotational events. These newly observed states fit with F₁ structures that were inconsistent with the canonical mechanism, and indicate that catalytic dwell oscillations must persist until the correct occupancy of substrates and products occurs at all three catalytic sites. When that condition is met, F₁ can proceed to the next power stroke. Understanding the basis of these catalytic dwell oscillations completes the F₁-ATPase rotary mechanism.