Johnson K A
J Biol Chem. 1983 Nov 25;258(22):13825-32.
The kinetics of ATP binding and hydrolysis (formation of acid-labile phosphate) by the Tetrahymena 30 S dynein ATPase has been measured by chemical quench flow methods. The amplitude of the ATP-binding transient gave a molecular weight per ATP-binding site of approximately 750,000, suggesting nearly 3 ATP binding sites/2 million Mr dynein molecule (Johnson, K. A., and Wall, J.S. (1983) J. Cell Biol. 96, 669-678). ATP binding occurred at the rate predicted from the apparent second order rate constant of 4.7 X 10(6) M-1 S-1 measured by analysis of the ATP-induced dissociation of the microtubule-dynein complex (Porter, M. E., and Johnson, K. A. (1983) J. Biol. Chem. 258, 6582-6587). Hydrolysis was slower than binding and occurred at a rate of 55 S-1, at 30 and 50 microM ATP. The rate limiting step for steady state turnover (product release) occurred with a rate constant of 8 S-1. These data show that the first two steps of the pathway of coupling ATP hydrolysis to the microtubule-dynein cross-bridge cycle are the same as those described by Lymn and Taylor for actomyosin (Lymn, R. W., and Taylor, E. W. (1971) Biochemistry 10, 4617-4624). Namely, ATP binding induces the very rapid dissociation of dynein from the microtubule and ATP hydrolysis occurs more slowly following dissociation. Moreover, in spite of rather gross structural differences, the kinetic constants for dynein and myosin are quite similar.
利用化学淬灭流动法测定了嗜热四膜虫30S动力蛋白ATP酶与ATP结合及水解（形成酸不稳定磷酸盐）的动力学。ATP结合瞬变的幅度给出每个ATP结合位点的分子量约为750,000，表明每200万道尔顿的动力蛋白分子中约有3个ATP结合位点（约翰逊，K.A.，和沃尔，J.S.（1983年）《细胞生物学杂志》96卷，669 - 678页）。ATP结合速率是根据通过分析ATP诱导的微管 - 动力蛋白复合物解离测得的表观二级速率常数4.7×10⁶ M⁻¹ s⁻¹预测得出的（波特，M.E.，和约翰逊，K.A.（1983年）《生物化学杂志》258卷，6582 - 6587页）。在30和50微摩尔ATP浓度下，水解比结合慢，水解速率为55 s⁻¹。稳态周转（产物释放）的限速步骤以8 s⁻¹的速率常数发生。这些数据表明，将ATP水解与微管 - 动力蛋白横桥循环偶联的途径的前两个步骤与Lymn和Taylor描述的肌动球蛋白的步骤相同（Lymn，R.W.，和Taylor，E.W.（1971年）《生物化学》10卷，4617 - 4624页）。即，ATP结合诱导动力蛋白从微管上非常快速地解离，并且在解离后ATP水解发生得更慢。此外，尽管存在相当大的结构差异，但动力蛋白和肌球蛋白的动力学常数相当相似。
