The magnesium ion-dependent adenosine triphosphatase of myosin. Two-step processes of adenosine triphosphate association and adenosine diphosphate dissociation.

The kinetics of protein-fluorescence change when rabbit skeletal myosin subfragment 1 is mixed with ATP or adenosine 5'-(3-thiotriphosphate) in the presence of Mg(2+) are incompatible with a simple bimolecular association process. A substrate-induced conformation change with DeltaG(0)<-24kJ.mol(-1) (i.e. DeltaG(0) could be more negative) at pH8 and 21 degrees C is proposed as the additional step in the binding of ATP. The postulated binding mechanism is M+ATPright harpoon over left harpoonM.ATPright harpoon over left harpoonM*.ATP, where the association constant for the first step, K(1), is 4.5x10(3)m(-1) at I 0.14m and the rate of isomerization is 400s(-1). In the presence of Mg(2+), ADP binds in a similar fashion to ATP, the rate of the conformation change also being 400s(-1), but with DeltaG(0) for that process being -14kJ.mol(-1). The effect of increasing ionic strength is to decrease K(1), the kinetics of the conformation change being essentially unaltered. Alternative schemes involving a two-step binding process for ATP to subfragment 1 are possible. These are not excluded by the experimental results, although they are perhaps less likely because they imply uncharacteristically slow bimolecular association rate constants.