Catalysis and rotation of F1 motor: cleavage of ATP at the catalytic site occurs in 1 ms before 40 degree substep rotation.
F1, a water-soluble portion of FoF1-ATP synthase, is an ATP hydrolysis-driven rotary motor. The central gamma-subunit rotates in the alpha 3 beta 3 cylinder by repeating the following four stages of rotation: ATP-binding dwell, rapid 80 degrees substep rotation, interim dwell, and rapid 40 degrees substep rotation. At least two 1-ms catalytic events occur in the interim dwell, but it is still unclear which steps in the ATPase cycle, except for ATP binding, correspond to these events. To discover which steps, we analyzed rotations of F1 subcomplex (alpha 3 beta 3 gamma) from thermophilic Bacillus PS3 under conditions where cleavage of ATP at the catalytic site is decelerated: hydrolysis of ATP by the catalytic-site mutant F1 and hydrolysis of a slowly hydrolyzable substrate ATP gamma S (adenosine 5'-[gamma-thio]triphosphate) by wild-type F1. In both cases, interim dwells were extended as expected from bulk phase kinetics, confirming that cleavage of ATP takes place during the interim dwell. Furthermore, the results of ATP gamma S hydrolysis by the mutant F1 ensure that cleavage of ATP most likely corresponds to one of the two 1-ms events and not some other faster undetected event. Thus, cleavage of ATP on F1 occurs in 1 ms during the interim dwell, and we call this interim dwell catalytic dwell.
Shimabukuro
Katsuya
K
Chemical Resources Laboratory, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8503, Japan.
Yasuda
Ryohei
R
Muneyuki
Eiro
E
Hara
Kiyotaka Y
KY
Kinosita
Kazuhiko
K
Yoshida
Masasuke
M
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D001407:Bacillus / Q000201:enzymology; D001665:Binding Sites; D001671:Biochemistry / Q000379:methods; D002384:Catalysis; D020134:Catalytic Domain; D004305:Dose-Response Relationship, Drug; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D020409:Molecular Motor Proteins; D009154:Mutation; D011485:Protein Binding; D006180:Proton-Translocating ATPases / Q000737:chemistry*; D012399:Rotation; D013997:Time Factors
D020409:Molecular Motor Proteins; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases