Studies on the actin activation of myosin subfragment-1 isoezymes and the role of myosin light chains.
Studies on the actin activation of myosin subfragment-1 isoezymes and the role of myosin light chains.
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肌球蛋白亚片段1同工酶的肌动蛋白激活及肌球蛋白轻链作用的研究。
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发表时间：
1979
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通讯作者：
A. Weeds
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作者：
Paul D. Wagner;Cecilia S. Slater;B. Pope;A. Weeds
肌动蛋白激活的肌球蛋白亚片段1 （S-1）同工酶的atp酶活性，根据其碱轻链（A1和A2）含量进行了分离，在许多不同的条件下进行了分析。先前关于肌动蛋白浓度增加对ATP酶影响的实验表明，S-1 （A2）对ATP的最大周转率大约是S-1 （A1）的两倍，肌动蛋白的Km也相当大。在这里，我们表明当使用调节的肌动蛋白（肌动蛋白+原肌球蛋白+肌钙蛋白）代替纯化的f -肌动蛋白时，这些动力学差异得以维持。然而，将溶液的离子强度从6 mM增加到46 mM KCl对S-1 （A2）的V影响不大，但S-1 （A1）的相应值增加到与S-1 （A2）大致相等，这表明在这些条件下，两种同工酶在稳态下受到相同的限速过程控制。两种同工酶的Km值也随离子强度的增加而增加，但S-1 （A1）的Km值增加得更快，接近S-1 （A2）。这些实验都是在固定的S-1浓度下进行的，但在固定的肌动蛋白浓度和S-1浓度变化的实验中，也可以得到S-1 atp酶的最大肌动蛋白活化速率。在6 mM KCl条件下，无限S-1浓度下S-1 （A1）和S-1 （A2）每摩尔肌动蛋白水解ATP的最大速率是相同的，这个数值等于无限S-1浓度下S-1 （A2）水解ATP的最大速率。这些结果表明，S-1 （A2）在atp酶循环中的限速步骤是肌动蛋白·S-1·ADP·Pi络合物的产物释放，而S-1 （A1）在无限肌动蛋白浓度下的V值相当低，这一过程可能发生在肌动蛋白与S-1·ADP·Pi络合物重新结合之前，使用lynn - taylor动力学模型。因此，增加离子强度似乎改变了actoS-1稳态水解ATP的限速过程（A1）。这些实验讨论了碱轻链在肌球蛋白atp酶和肌肉收缩中的作用。
The actin-activated ATPase activity of myosin subfragment 1 (S-1) isoenzymes, separated on the basis of their alkali light chain (A1 and A2) content, has been analysed under a number of different conditions. Previous experiments on the effects of increasing actin concentrations on the ATPase have shown that the maximum turnover rate of ATP by S-1 (A2) was about twice that of S-1 (A1), and the Km for actin was also considerably larger for this isoenzyme. Here we show that these kinetic differences are maintained when regulated actin (actin + tropomyosin + troponin) is used in place of purified F-actin. However, while increasing the ionic strength of the solution from 6 mM to 46 mM KCl has little effect on V for S-1 (A2), the corresponding value for S-1 (A1) increases to approximately equal that of S-1 (A2), suggesting that the two isoenzymes are controlled by the same rate-limiting process in the steady state under these conditions. The value of Km also increases with increasing ionic strength for the two isoenzymes, but that for S-1 (A1) increases more rapidly and approaches that for S-1 (A2). These experiments were carried out at at fixed S-1 concentration, but the maximum rate of actin activation of the S-1 ATPase can also be obtained from experiments where the actin concentration is fixed and the S-1 concentration varied. Under these conditions in 6 mM KCl, the maximum rate of ATP hydrolysis expressed per mole of actin at infinite S-1 concentration is the same for both S-1 (A1) and S-1 (A2), and this value numerically equals the maximum rate of hydrolysis of ATP by S-1 (A2) expressed at infinite actin concentration. These results suggest that the rate-limiting step in the ATPase cycle for S-1 (A2) is the release of products from an actin · S-1 · ADP · Pi complex, while for S-1 (A1), where the value of V at infinite actin concentration is considerably lower, a different process is rate limiting, and this process may occur before actin reassociation with the S-1 · ADP · pi complex, using the Lymn-Taylor kinetic model. Thus increasing the ionic strength appears to change the rate-limiting process in the steady-state hydrolysis of ATP by actoS-1 (A1). These experiments are discussed in terms of the role of the alkali light chains in the myosin ATPase and muscle contraction.