Catalytic dwell oscillations complete the F1-ATPase mechanism

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. 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. However, the complete order of catalytic events is not fully understood. Here, the authors monitor the subunit-γ rotary position every 5 μs for successive 0.125 ms intervals, revealing catalytic dwell oscillations in three successive stages as a key aspect of completing the F1-ATPase rotary mechanism.