Inhibition of thermophilic F(1)-ATPase by the ε subunit takes different path from the ADP-Mg inhibition

The F(1)-ATPase, the soluble part of FoF(1)-ATP synthase, is a rotary molecular motor consisting of α(3)β(3)γδε. The γ and ε subunits rotate relative to the α(3)β(3)δ sub-complex on ATP hydrolysis by the β subunit. The ε subunit is known as an endogenous inhibitor of the ATPase activity of the F(1)-ATPase and is believed to function as a regulator of the ATP synthase. This inhibition by the ε subunit (ε inhibition) of F(1)-ATPase from thermophilic Bacillus PS3 was analyzed by single molecule measurements. By using a mutant ε subunit deficient in ATP binding, reversible transitions between active and inactive states were observed. Analysis of pause and rotation durations showed that the ε inhibition takes a different path from the ADP-Mg inhibition. Furthermore, the addition of the mutant ε subunit to the α(3)β(3)γ sub-complex was found to facilitate recovery of the ATPase activity from the ADP-Mg inhibition. Thus, it was concluded that these two inhibitions are essentially exclusive of each other.