F1FO ATP synthase molecular motor mechanisms

Abstract

The F-ATP synthase, consisting of F₁ and F_O motors connected by a central rotor and the stators, is the enzyme responsible for synthesizing the majority of ATP in all organisms. The F₁ (αβ)₃ ring stator contains three catalytic sites. Single-molecule F₁ rotation studies revealed that ATP hydrolysis at each catalytic site (0°) precedes a power-stroke that rotates subunit-γ 120° with angular velocities that vary with rotational position. Catalytic site conformations vary relative to subunit-γ position (β_E, empty; β_D, ADP bound; β_T, ATP-bound). During a power stroke, β_E binds ATP (0°-60°) and β_D releases ADP (60°-120°). Årrhenius analysis of the power stroke revealed that elastic energy powers rotation via unwinding the γ-subunit coiled-coil. Energy from ATP binding at 34° closes β_E upon subunit-γ to drive rotation to 120° and forcing the subunit-γ to exchange its tether from β_E to β_D, which changes catalytic site conformations. In F₁F_O, the membrane-bound F_O complex contains a ring of c-subunits that is attached to subunit-γ. This c-ring rotates relative to the subunit-a stator in response to transmembrane proton flow driven by a pH gradient, which drives subunit-γ rotation in the opposite direction to force ATP synthesis in F₁. Single-molecule studies of F₁F_O embedded in lipid bilayer nanodisks showed that the c-ring transiently stopped F₁-ATPase-driven rotation every 36° (at each c-subunit in the c₁₀-ring of E. coli F₁F_O) and was able to rotate 11° in the direction of ATP synthesis. Protonation and deprotonation of the conserved carboxyl group on each c-subunit is facilitated by separate groups of subunit-a residues, which were determined to have different pKa's. Mutations of any of any residue from either group changed both pKa values, which changed the occurrence of the 11° rotation proportionately. This supports a Grotthuss mechanism for proton translocation and indicates that proton translocation occurs during the 11° steps. This is consistent with a mechanism in which each 36° of rotation the c-ring during ATP synthesis involves a proton translocation-dependent 11° rotation of the c-ring, followed by a 25° rotation driven by electrostatic interaction of the negatively charged unprotonated carboxyl group to the positively charged essential arginine in subunit-a.

Keywords: F1 ATPase; F1Fo ATP synthase; rotary molecular motor; single-molecule studies; torque.