Redox Regulation of Rotation of the Cyanobacterial F1-ATPase Containing Thiol Regulation Switch

F-1-ATP synthase (F-1-ATPase) is equipped with a special mechanism that prevents the wasteful reverse reaction, ATP hydrolysis, when there is insufficient proton motive force to drive ATP synthesis. Chloroplast F-1-ATPase is subject to redox regulation, whereby ATP hydrolysis activity is regulated by formation and reduction of the disulfide bond located on the gamma subunit. To understand the molecular mechanism of this redox regulation, we constructed a chimeric F-1 complex (alpha(3)beta(3)gamma(redox)) using cyanobacterial F-1, which mimics the regulatory properties of the chloroplast F-1-ATPase, allowing the study of its regulation at the single molecule level. The redox state of the gamma subunit did not affect the ATP binding rate to the catalytic site(s) and the torque for rotation. However, the long pauses caused by ADP inhibition were frequently observed in the oxidized state. In addition, the duration of continuous rotation was relatively shorter in the oxidized alpha(3)beta(3)gamma(redox) complex. These findings lead us to conclude that redox regulation of CF1-ATPase is achieved by controlling the probability of ADP inhibition via the gamma subunit inserted region, a sequence feature observed in both cyanobacterial and chloroplast ATPase gamma subunits, which is important for ADP inhibition (Sunamura, E., Konno, H., Imashimizu-Kobayashi, M., Sugano, Y., and Hisabori, T. (2010) Plant Cell Physiol. 51, 855-865).