pH-dependent 11° F ₁ F _o ATP synthase sub-steps reveal insight into the F _o torque generating mechanism

Most cellular ATP is made by rotary F ₁ F _o ATP synthases using proton translocation-generated clockwise torque on the F _o c-ring rotor, while F ₁ -ATP hydrolysis can force anticlockwise rotation and proton pumping. Although the interface of stator subunit-a containing the transmembrane half-channels and the c-ring is known from recent F ₁ F _o structures, the torque generating mechanism remains elusive. Here, single-molecule studies reveal pH-dependent 11° rotational sub-steps in the ATP synthase direction of the E. coli c ₁₀ -ring of F ₁ F _o against the force of F ₁ -ATPase-dependent rotation that result from H ⁺ transfer events from F _o subunit-a groups with a low pKa to one c-subunit of the c-ring, and from an adjacent c-subunit to stator groups with a high pKa. Mutations of subunit-a residues in the proton translocation channels alter these pKa values, and the ability of synthase substeps to occur. Alternating 11° and 25° sub-steps then result in sustained ATP synthase rotation of the c ₁₀ -ring.