Biophysical Characterization of a Thermoalkaliphilic Molecular Motor with a High Stepping Torque Gives Insight into Evolutionary ATP Synthase Adaptation.
F1F0ATP synthases are bidirectional molecular motors that translocate protons across the cell membrane by either synthesizing or hydrolyzing ATP. Alkaliphile ATP synthases are highly adapted, performing oxidative phosphorylation at high pH against an inverted pH gradient (acidin/alkalineout). Unlike mesophilic ATP synthases, alkaliphilic enzymes have tightly regulated ATP hydrolysis activity, which can be relieved in the presence of lauryldimethylamine oxide. Here, we characterized the rotary dynamics of the Caldalkalibacillus thermarum TA2.A1 F1ATPase (TA2F1) with two forms of single molecule analysis, a magnetic bead duplex and a gold nanoparticle. TA2F1rotated in a counterclockwise direction in both systems, adhering to Michaelis-Menten kinetics with a maximum rotation rate (Vmax) of 112.4 revolutions/s. TA2F1displayed 120° unitary steps coupled with ATP hydrolysis. Torque measurements revealed the highest torque (52.4 piconewtons) derived from an F1molecule using fluctuation theorem. The implications of high torque in terms of extreme environment adaptation are discussed.
From the Department of Applied Chemistry, Graduate School of Engineering, The University of Tokyo, Tokyo 113-8656, Japan and duncan.mcmillan@nojilab.t.u-tokyo.ac.jp.
Duncan G G
DG
McMillan
From the Department of Applied Chemistry, Graduate School of Engineering, The University of Tokyo, Tokyo 113-8656, Japan and.
Rikiya
R
Watanabe
From the Department of Applied Chemistry, Graduate School of Engineering, The University of Tokyo, Tokyo 113-8656, Japan and.
Hiroshi
H
Ueno
the Department of Microbiology and Immunology, Otago School of Medical Sciences, University of Otago, P. O. Box 56, Dunedin 9054, New Zealand.
Gregory M
GM
Cook
From the Department of Applied Chemistry, Graduate School of Engineering, The University of Tokyo, Tokyo 113-8656, Japan and hnoji@appchem.t.u-tokyo.ac.jp.
Hiroyuki
H
Noji
D001406:Bacillaceae / Q000201:enzymology* / Q000235:genetics; D004123:Dimethylamines / Q000737:chemistry; D019143:Evolution, Molecular; D007700:Kinetics; D008958:Models, Molecular; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D004123:Dimethylamines; C014518:dodecyldimethylamine oxide; D006180:Proton-Translocating ATPases
ATP synthase; F1F0-ATPase; adaptation; enzyme kinetics; enzyme mechanism; respiratory chain; rotation; single molecule biophysics; thermoalkaliphile; torque