The concerted nature between three catalytic subunits driving the F1 rotary motor

F1, a rotational molecular motor, shows strong cooperativity during ATP catalysis when driving the rotation of the central γ subunit surrounded by the α3β3 subunits. To understand how the three catalytic β subunits cooperate to drive rotation, we made a hybrid F1 containing one or two mutant β subunits with altered catalytic kinetics and observed its rotations. Analysis of the asymmetric stepwise rotations elucidated a concerted nature inside the F1 complex where all three β subunits participate to rotate the γ subunit with a 120° phase. In addition, observing hybrid F1 rotations at various solution conditions, such as ADP, Pi and the ATPase inhibitor 2,3-butanedione 2-monoxime (BDM) provides additional information for each elementary event. This novel experimental system, which combines single molecule observations and biochemical methods, enables us to dynamically visualize the catalytic coordination inside active enzymes and shed light on how biological machines provide unidirectional functions and rectify information from stochastic reactions.