F-1-ATPase (F-1), an important rotary motor protein, converts the chemical energy of ATP hydrolysis into mechanical energy using rotary motion with extremely high efficiency. The energy-conversion mechanism for this molecular motor has been extensively clarified by previous studies, which indicate that the interactions between the catalytic residues and the beta- and gamma-phosphates of ATP are indispensable for efficient catalysis and torque generation. However, the role of alpha-phosphate is largely unknown. In this study, we observed the rotation of F-1 fuelled with an ATP analogue, adenosine 5'-[alpha-thio]-triphosphate (ATP alpha S), in which the oxygen has been substituted with a sulfur ion to perturb the alpha-phosphate/F-1 interactions. In doing so, we have revealed that ATPaS does not appear to have any impact on the kinetic properties of the motor or on torque generation compared to ATP. On the other hand, F-1 was observed to lapse into the ADP-inhibited intermediate states when in the presence of ATPaS more severely than in the presence of ATP, suggesting that the alpha-phosphate group of ATP contributes to the avoidance of ADP-inhibited intermediate formation. (C) 2015 Elsevier Inc. All rights reserved.