F1FO ATP synthase molecular motor mechanisms

Abstract

The F-ATP synthase, consisting of F1 and FO 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 (βE, empty; βD, ADP bound; βT, ATP-bound). During a power stroke, βE binds ATP (0°–60°) and βD 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 βE upon subunit-γ to drive rotation to 120° and forcing the subunit-γ to exchange its tether from βE to βD, which changes catalytic site conformations. In F1FO, the membrane-bound FO 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 F1FO 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 F1FO) 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.

Changes within the central stalk of E. coli F1Fo ATP synthase observed after addition of ATP

“…2c ) and bind MgADP. Comparison of the relative position of the rotor axel between known structures 8 , 16 , 19 , 26 – 28 , highlighted that the F 1 -ATPase was in a similar rotary position to that of observed of Geobacillus stearothermophilus (also termed Bacillus PS3) in the catalytic dwell 8 (Supplementary Fig. 3 ), indicating that this enzyme was in a similar state.…”

Rotary mechanism of V/A-ATPases—how is ATP hydrolysis converted into a mechanical step rotation in rotary ATPases?

“…ATP synthases are enzymes that play crucial roles in energy metabolism ( Boyer, 1997 ; Yoshida et al, 2001 ; Kuhlbrandt, 2019 ; Frasch et al, 2022 ). They convert an electrochemical proton motive force across the membrane generated by respiration into chemical energy that is stored in the form of ATP.…”

“…ATP synthases are classified into two major categories: F-type ATP synthases (F o F 1 -ATPase) and V-type ATP synthases (V/A-ATPase) ( Figures 1A, B ) ( Yokoyama and Imamura, 2005 ; Forgac, 2007 ; Vasanthakumar and Rubinstein, 2020 ). F o F 1 -ATPases are found in the inner membrane of mitochondria, the thylakoid membrane of chloroplasts, and the plasma membrane of eubacteria ( Frasch et al, 2022 ). V/A-ATPases are present in some eubacteria and archaea and are structurally similar to eukaryotic V-ATPases.…”

Single-molecule FRET dynamics of molecular motors in an anti-Brownian electrokinetic trap

“…(The structural details and functional mechanism are reviewed elsewhere. [19][20][21][22][23][24] ) The 3-fold symmetry of the head is reflected by the 120°-stepping motion of the rotor domain. It can be visualised by labelling the rotor and stator subunits with a pair of fluorescent dyes and monitoring the periodic intensity fluctuations of the donor and acceptor dye during rotation by smFRET after reconstitution of a single F O F 1 -ATP synthase in an energised liposome.…”