[Affinity modification of myosin with protected active centers: confirmation of the existence of an allosteric substrate-binding segment].

The effect of an affinity modifier of myosin ATPase representing a mixed anhydride of AMP and mesitylene carboxylic acid (AMP-MA) on myosin with protected active centers was studied. The protection of active centers was performed by the method of Wells et al. Which consists in the stabilization of the myosin-MgADP complex in the enzyme active center by way of cross-linking of the active center with a Co-phenanthroline complex simultaneously interacting with two SH-groups of the protein. Myosin with protected active center completely loses its ability to hydrolyze ATP; however, it can be reactivated by way of SH-group reduction with a subsequent MgADP release from the active centers. Treatment of myosin with protected active centers with AMP-MA does not result in the reduction of the enzyme activity after removal of the Co-phenanthroline complex. This suggests that the irreversible inhibition of myosin ATPase by AMP-MA occurs due to the protein modification outside the active center(s), which provides support for our earlier made conclusion concerning the existence of an additional (with respect to active centers) substrate-binding site in the myosin molecule.