Oxygen exchange during the acto-subfragment-1 ATPase reaction: evidence for the two-route mechanism of the actomyosin ATPase reaction.
The oxygen exchange occurring during the acto-S-1 ATPase reaction was analyzed based on the distribution of 18O-labeled species of P1 using [gamma-18O]ATP as a substrate. Evidence was found for the two-route mechanism in which ATP is hydrolyzed via the dissociation of acto-S-1 into F-actin and the S-1-phosphate-ADP complex, S-1PADP, and their recombination, and also hydrolyzed without the dissociation of acto-S-1 (Inoue, A., Shigekawa, M., & Tonomura, Y. (1973) J. Biochem. 74, 923-934; Inoue, A., Ikebe, M., & Tonomura, Y. (1980) J. Biochem. 88, 1663-1677). When ATP was mainly hydrolyzed without the dissociation of acto-S-1, the extent of oxygen exchange was low. When ATP was hydrolyzed by both routes, the distribution of product P1 with 3, 2, 1, and 0 18O atoms showed a mixture resulting from low and high oxygen exchange. The rate of ATPase without the dissociation of acto-S-1 can be estimated from the rate of the overall reaction (v), the rate of recombination of S-1PADP with F-actin (vr), and the extent of dissociation of acto-S-1 (a). The distribution of the P1 species measured was almost equal to that calculated from the ratio of ATP hydrolysis via the two pathways as avr and v-avr, respectively. This result indicates that the rates of the dissociation of acto-S-1PADP into S-1PADP and F-actin and their recombination are much lower than the rate of decomposition of the acto-S-1PADP complex into acto-S-1 + ADP + Pi.
Yasui
M
M
Department of Biology, Faculty of Science, Osaka University.
Ohe
M
M
Kajita
A
A
Arata
T
T
Inoue
A
A
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D006868:Hydrolysis; D007553:Isotope Labeling / Q000379:methods; D007700:Kinetics; D009132:Muscles / Q000201:enzymology*; D015879:Myosin Subfragments; D009218:Myosins / Q000378:metabolism*; D010103:Oxygen Isotopes; D010446:Peptide Fragments / Q000378:metabolism*; D051381:Rats
D015879:Myosin Subfragments; D010103:Oxygen Isotopes; D010446:Peptide Fragments; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D009218:Myosins