The mechanism of ATP hydrolysis catalyzed by myosin and actomyosin, using rapid reaction techniques to study oxygen exchange.
During ATP hydrolysis, myosin or subfragment 1 catalyzes the exchange of oxygens between water and phosphate, so that on average, each product Pi molecule contains more than one oxygen atom derived from water. Using quenched-flow techniques, the exchange process in both ATP and Pi was studied in the rabbit skeletal muscle subfragment 1 ATPase. The exchange in protein-bound ATP (M*.ATP) and protein-bound ADP.Pi (M**.ADP.Pi) was followed as a function of time. The pattern of exchange follows closely a model in which M*.ATP + H2O and M**.ADP.Pi are interconverted directly, without any characterizable intermediate between these species. All oxygen atoms of Pi are equivalent with respect to loss to solvent, and elimination of water to re-form ATP occurs with a rate constant of 15 s-1 (at pH 8.0, I 0.015 M, 20 degrees C). This is the first direct measurement of the rate constant for the transformation of M**.ADP.Pi to M*.ATP. Oxygen exchange during steady state ATP hydrolysis in the presence of acto-subfragment 1 also fits well to this model, with actin reducing the time available for M*.ATP and M**.ADP.Pi to undergo exchange.
D000205:Actomyosin / Q000378:metabolism*; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D007700:Kinetics; D009132:Muscles / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D010103:Oxygen Isotopes; D010710:Phosphates; D011817:Rabbits
D010103:Oxygen Isotopes; D010710:Phosphates; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000205:Actomyosin; D009218:Myosins