pH-dependent 11° F₁F_o ATP synthase sub-steps reveal insight into the F_o torque generating mechanism

Most cellular ATP is made by rotary F₁F_o ATP synthases using proton translocation-generated clockwise torque on the F_o c-ring rotor, while F₁-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 F₁F_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 c₁₀-ring of F₁F_o against the force of F₁-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 c₁₀-ring.