Interaction of ADP with Skeletal and Cardiac Myosin and Their Active Fragments Observed by Proton Release

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Abstract
The technique of proton release measurement has been used to explore the binding of ADP to skeletal and cardiac myosins and their active fragments in a variety of conditions. Binding profiles were obtained on intact myosin in the filamentous, undissolved form in physiological solvent conditions. Binding constants are given. At higher ionic strength (0.5 M KCl) the binding profile of Mg ADP is compatible with the presence of 2 types of site, differing from one another both in respect of affinity and the number of protons released per site. Studies with cardiac myosin reveal no such indications of heterogeneity and are consistent with the presence of a single population of thermodynamically indistinguishable sites. In the absence of divalent cations, in solutions containing K ions and EDTA, ADP binds with absorption rather than liberation of protons. The pH profile of proton absorption at saturation can be fitted in terms of an ionizing group with an unperturbed pK of 9.4, and at least 1 of lower pK (5.9). The dissociation constant (pH 8 at 5.degree. C) is about 8 .mu.M, and the affinity for uncomplexed ADP is thus only slightly weaker than that for Mg ADP.