Cooperation amongc-subunits of F_oF₁-ATP synthase in rotation-coupled proton translocation

Abstract

In FoF1-ATP synthase, proton translocation through Fo drives rotation of the c-subunit oligomeric ring relative to the a-subunit. Recent studies suggest that in each step of the rotation, key glutamic acid residues in different c-subunits contribute to proton release to and proton uptake from the a-subunit. However, no studies have demonstrated cooperativity among c-subunits toward FoF1-ATP synthase activity. Here, we addressed this using Bacillus PS3 ATP synthase harboring c-ring with various combinations of wild-type and cE56D, enabled by genetically fused single-chain c-ring. ATP synthesis and proton pump activities were significantly decreased by a single cE56D mutation and further decreased by double cE56D mutations. Moreover, activity further decreased as the two mutation sites were separated, indicating cooperation among c-subunits. Similar results were obtained for proton transfer-coupled molecular simulations. Simulations revealed that prolonged proton uptake in mutated c-subunits is shared between two c-subunits, explaining the cooperation observed in biochemical assays.

Regulatory Mechanisms and Environmental Adaptation of the F-ATPase Family

“…31,32) The H + pathway of F O , which is formed from two half channels that open on each side of the membrane, consists of the a subunit and the c-ring-composed of 8-15 c subunits, varying in number among species. [33][34][35][36] The c subunit has an essential glutamic acid (Glu) or aspartic acid (Asp) residue that binds H + from one half of the channel and releases it into the other one, leading to c-ring rotation [37][38][39] (Fig. 1).…”