Kinetic Analysis of the Myosin B-Adenosine-Triphosphatase System*

Abstract

Since Engelhardt and Ljubimova (1) found the ATPase activity of actomyosin and myosin, its role in muscle contraction has become the subjects of repeated investigations (2, 3). The general properties of this enzyme were elucidated by Banga (4), Mommaerts and Seradarian (5) and Hasselbach (6). These authors showed that Ca⁺⁺ accelerates ATPase under all conditions investigated, while Mg⁺⁺ inhibits ATPase of actomyosin at high KC1 concentration and accelerates it at low KC1 and also that the pH-activity curve has a characteristic shape showing a maximum and a minimum at pH about 6.5 and 7.5, respectively. More recently, several authors have demonstrated that myosin catalyses the hydrolysis of ITP, GTP, UTP and CTP (7–11) in addition to ATP and that it is activated markedly by EDTA (12–14), PP (15, 16), AET (17), PCMB (18) and DNP (19, 20). Ouellet et al. (21), Tonomura and Watanabe (22, 23) and Green and Mommaerts (24) have already found that ATPase action obeys, in various ionic media, the Michaelis-Menten theory. These kinetic studies have elucidated the mechanism of this enzyme action to some extent. A thorough kinetic study, however, is needed in order to clarify the physico-chemical mechanism of the effect of divalent cations, pH and the particular activators. In view of these circumstances, the present authors have reinvestigated the kinetic properties of myosin B-ATPase at the steady state** in various ionic media and obtained the following results: (i) Ca⁺⁺ activates ATPase by the chelation of ATP with Ca⁺⁺, (ii) the effects of divalent cations depend mainly on their ionic radii, (iii) at sufficiently high concentration of the substrate, ATPase is activated again by the substrate, (iv) in the presence of high concentrations of the substrate or EDTA, the pH-activity curve lacks the characteristic depression at pH about 7.5, and (v) ITPase bears some resemblance to ATPase in the presence of the particular activator. These results have enabled us to propose a general theory of the reaction mechanism of myosin B-ATPase.