Search

The Michaelis constant ( K m ) and the maximum velocity ( v m ) of myosin A-ATPase (EC 3.6.1.3) and of myosin A-ITPase were measured in 0.6 M KCl, and in the presence of 7 mM CaCl 2 , over a wide range of temperature and pH. The following results were obtained: 1. 1. Both v m and K m of ITPase were increased proportionally to each other with increasing pH, and their half-saturation values were obtained at pH 6.8. The Arrhenius plots of v m and K m were readily represented by two reactions of different activation energies and the extrapolations of the linear portions intersected in each case at around 15°. The value of activation energies were 25.6 and 11.9 kcal/mole for v m and 7.9 and 2.1 kcal/mole for K m below and above 15°, respectively. 2. 2. At 20°, the K m of ATPase changed with changing pH in proportion to v m : both K m and v m showed a depression at neutral pH. The pH dependence of the ratio of v m of ITPase to that of ATPase was given by a bell-shaped curve having its maximum at pH 7.5. At 0°, the pH-activity curve of ATPase lacked the neutral depression. At pH 6, the Arrhenius plots of v m and K m were linear and the activation energies of v m and K m were 16.2 and −2.8 kcal/mole, respectively. At pH7, the activation energies of v m and K m , at temperatures higher than 10°, were noticeably different from those at temperatures lower than 10°; these values were for v m , 9.0 and 3.0 kcal/mole, and for K m , −1.9 and −10.4 kcal/mole, below and above 10°, respectively. 3. 3. These results were analyzed according to the mechanism proposed by one of the authors where by, in ATPase, two forms of the Michaelis complex, viz. , an active one (E 1 S) and an inactive one (E 2 S), can exist. It was concluded that the complex E 2 S is not formed in ITPase but in ATPase it is dominant at around pH 7.5, and at higher temperatures. It was further concluded that the rate-constant of dissociation of the Michaelis complex E 1 S into the enzyme and the substrate is much smaller than that of the breakdown of the complex into the products; the rate-constant of the formation of E 1 S is independent of pH.