Power Stroke Angular Velocity Profiles of Archaeal A-ATP Synthase Versus Thermophilic and Mesophilic F-ATP Synthase Molecular Motors

Abstract

Edited by Velia FowlerThe angular velocities of ATPase-dependent power strokes as a function of the rotational position for the A-type molecular motor A 3 B 3 DF, from the Methanosarcina mazei Gö1 A-ATP synthase, and the thermophilic motor ␣ 3 ␤ 3 ␥, from Geobacillus stearothermophilus (formerly known as Bacillus PS3) F-ATP synthase, are resolved at 5 s resolution for the first time. Unexpectedly, the angular velocity profile of the A-type was closely similar in the angular positions of accelerations and decelerations to the profiles of the evolutionarily distant F-type motors of thermophilic and mesophilic origins, and they differ only in the magnitude of their velocities. M. mazei A 3 B 3 DF power strokes occurred in 120°steps at saturating ATP concentrations like the F-type motors. However, because ATP-binding dwells did not interrupt the 120°steps at limiting ATP, ATP binding to A 3 B 3 DF must occur during the catalytic dwell. Elevated concentrations of ADP did not increase dwells occurring 40°after the catalytic dwell. In F-type motors, elevated ADP induces dwells 40°after the catalytic dwell and slows the overall velocity. The similarities in these power stroke profiles are consistent with a common rotational mechanism for A-type and F-type rotary motors, in which the angular velocity is limited by the rotary position at which ATP binding occurs and by the drag imposed on the axle as it rotates within the ring of stator subunits.The primary source of ATP in archaea such as methanogens is the A 1 A O -ATP synthase, which shares several structural features with eukaryotic V 1 V O -ATPases and is evolutionarily distant from the F 1 F O -ATP synthases that fulfill this role in other bacteria and eukaryotes (1, 2). Metabolism in archaea is coupled to the generation of H ϩ and/or Na ϩ potentials across the membrane, both of which can provide the energy for ATP synthesis by the A 1 A O -ATP synthase. Whereas the F 1 F O -ATP synthases in prokaryotes and eukaryotes catalyze ATP synthesis at the expense of an electrochemical ion potential, the evolutionarily related V 1 V O -ATPases function as ATP-driven ion pumps and are unable to synthesize ATP under physiological conditions (3)(4)(5). Although the cellular function of archaeal ATP synthases is to synthesize ATP powered by a non-equilibrium electrochemical gradient, they also work as ATP-driven ion pumps to generate an ion gradient under fermentative conditions (6).The A-ATP synthases are composed of two opposed rotary motors as follows: an integral membrane A O complex (ac x ) involved in ion translocation, and a peripheral A 1 complex (A 3 B 3 CDF) that contains the catalytic sites for a total of nine different subunits (6). Both motors are connected by two peripheral stalks each composed of an EH heterodimer. These complexes share common features with bacterial

The uniqueness of subunit α of mycobacterial F-ATP synthases: An evolutionary variant for niche adaptation

“…Upon addition of 1 mM MgATP, sinusoidal variations of light intensities scattered from the nanorod were observed as a function of its position relative to the axis of polarization. At a sampling rate of 1 kHz, dwells in the rotation produced a peak in the intensity distribution, as reported previously for (30). A series of 36 histograms of light intensities was recorded that were acquired when the rotational position of the polarizer was advanced successively by 10°.…”

“…To overcome these limitations, we used a 40 ϫ 76-nm gold nanorod as a probe of rotation. The viscous drag on these nanorods is not rate-limiting to the power stroke of the Escherichia coli ␣ 3 ␤ 3 ␥ (Ec␣ 3 ␤ 3 ␥) (28,29), Gs␣ 3 ␤ 3 ␥, or the A 3 B 3 DF of the Methanosarcina mazei Gö1 A 1 A 0 -ATP synthase (MmA 3 B 3 DF) (30). The light scattered from the nanorod was detected by an avalanche photo diode (APD) after passing through a polarizing filter using dark-field microscopy ( Fig.…”

“…The angular velocity profile for a 120°power stroke of the central stalk was calculated from the sinusoidal light intensity changes recorded with 200 kHz using the arcsin 1 ⁄2 function (30). The three off-set sinusoidal curves like that shown in Fig.…”

Method to extract multiple states in F 1 -ATPase rotation experiments from jump distributions

“…2A. They were extracted from the transitions using a method prompted by the work of Frasch and coworkers, (10,19), who use the "angular velocity" term for this normalized jump, ∆θ/∆t. In this work, θ is defined relative to the catalytic dwells clearly resolved in the trajectories (Fig.…”

Catalytic robustness and torque generation of the F1-ATPase

“…Because the observed profile is not consistent with any reported substep positions, a reasonable explanation for this conflict with previous reports is yet to be described. Sielaff (2016) reported a different torque profile. They observed that the instantaneous angular velocity of rotation probed with a gold nanorod showed a distinct peak during a 120° step.…”