Activation of pausing F1 motor by external force.
A rotary motor F(1), a catalytic part of ATP synthase, makes a 120 degrees step rotation driven by hydrolysis of one ATP, which consists of 80 degrees and 40 degrees substeps initiated by ATP binding and probably by ADP and/or P(i) dissociation, respectively. During active rotations, F(1) spontaneously fails in ADP release and pauses after a 80 degrees substep, which is called the ADP-inhibited form. In the present work, we found that, when pushed >+40 degrees with magnetic tweezers, the pausing F(1) resumes its active rotation after releasing inhibitory ADP. The rate constant of the mechanical activation exponentially increased with the pushed angle, implying that F(1) weakens the affinity of its catalytic site for ADP as the angle goes forward. This finding explains not only its unidirectional nature of rotation, but also its physiological function in ATP synthesis; it would readily bind ADP from solution when rotated backward by an F(o) motor in the ATP synthase. Furthermore, the mechanical work for the forced rotation was efficiently converted into work for expelling ADP from the catalytic site, supporting the tight coupling between the rotation and catalytic event.
Hirono-Hara
Yoko
Y
Institute of Industrial Science and Precursory Research for Embryonic Science and Technology, Japan Science and Technology Corporation, University of Tokyo, Tokyo 153-8505, Japan.
Ishizuka
Koji
K
Kinosita
Kazuhiko
K
Yoshida
Masasuke
M
Noji
Hiroyuki
H
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001407:Bacillus / Q000201:enzymology / Q000235:genetics; D025243:Bacterial Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001696:Biomechanical Phenomena; D055592:Biophysical Phenomena; D001703:Biophysics; D004789:Enzyme Activation; D008954:Models, Biological; D020409:Molecular Motor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics
D020409:Molecular Motor Proteins; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D025243:Bacterial Proton-Translocating ATPases