Sleep J A, Hutton R L
Biochemistry. 1978 Dec 12;17(25):5423-30. doi: 10.1021/bi00618a016.
The apparent second-order rate constant, ka-2, of actin binding to a myosin-ATP state (M*.ATP) and releasing ATP to the medium has been determined by two methods. The first was the measurement of the amount of ATP released when actin was added to the intermediate state, M*.ATP; the second was the measurement of oxygen exchange between ATP and HOH. A quantitative treatment of ATP in equilibrium HOH exchange is given to allow extraction of elementary rate constants from the data. Agreement between the two methods was good and at low ionic strength and 23 degrees C, ka-2 is 6 X 10(5) M-1 s-1 which is about one-third the value of the apparent second-order rate constant, ka4, of actin binding to the myosin product state (M**.ADP.Pi). The determination of ka-2 allows a lower limit of 6 s-1 to be placed upon the first-order rate of ATP release from AM.ATP. This is to be compared with a value of less than or equal to 1.5 X 10(-4) s-1 for the equivalent steps of the myosin scheme; thus actin enhances the rate by a factor of 4 X 10(4) or more. A greater proportion of the bound ATP is released to the medium as ATP with increasing actin concentration. This reflects the contribution to rate limitation at saturating actin concentration of steps between myosin states dissociated from actin.
肌动蛋白与肌球蛋白 - ATP 状态（M*.ATP）结合并将 ATP 释放到介质中的表观二级速率常数 ka-2 已通过两种方法测定。第一种方法是测量向中间状态 M*.ATP 添加肌动蛋白时释放的 ATP 量；第二种方法是测量 ATP 与 HOH 之间的氧交换。对平衡态 HOH 交换中的 ATP 进行了定量处理，以便从数据中提取基本速率常数。两种方法的结果吻合良好，在低离子强度和 23 摄氏度下，ka-2 为 6×10⁵ M⁻¹ s⁻¹，约为肌动蛋白与肌球蛋白产物状态（M**.ADP.Pi）结合的表观二级速率常数 ka4 值的三分之一。ka-2 的测定使得从 AM.ATP 释放 ATP 的一级速率下限为 6 s⁻¹。相比之下，肌球蛋白过程中同等步骤的值小于或等于 1.5×10⁻⁴ s⁻¹；因此，肌动蛋白将速率提高了 4×10⁴ 倍或更多。随着肌动蛋白浓度的增加，更大比例的结合 ATP 以 ATP 的形式释放到介质中。这反映了在饱和肌动蛋白浓度下，与肌动蛋白解离的肌球蛋白状态之间的步骤对速率限制的贡献。
