Hemric M E, Freedman M V, Chalovich J M
Department of Biochemistry, East Carolina University, School of Medicine, Greenville, North Carolina 27858-4354.
Arch Biochem Biophys. 1993 Oct;306(1):39-43. doi: 10.1006/abbi.1993.1477.
We have previously shown that caldesmon inhibits the actin-activated ATPase activity of myosin subfragments in parallel with inhibition of myosin subfragment.ATP binding to actin (M. E. Hemric, and J. M. Chalovich, 1988, J. Biol. Chem. 263, 1878-1885; L. Velaz, R. H. Ingraham, and J. M. Chalovich, 1990, J. Biol. Chem. 265, 2929-2934). From these data, we suggested that caldesmon is a competitive inhibitor of binding of myosin subfragment-1 to actin. To confirm this result, we now show the effect of caldesmon on the steady-state parameters of ATP hydrolysis by (A1)S-1 at increasing actin concentrations. Low ionic strength conditions were used to maximize the interaction between (A1)S-1 and actin. In both the presence and absence of smooth muscle tropomyosin, caldesmon caused a twofold decrease in the kcat and more than a 12-fold change in the KATPase. Therefore, competition of binding of myosin to actin by caldesmon contributes to the reduction in ATPase activity in both the presence and the absence of tropomyosin.
我们之前已经表明，钙调蛋白抑制肌球蛋白亚片段的肌动蛋白激活的ATP酶活性，同时抑制肌球蛋白亚片段与肌动蛋白的ATP结合（M.E. Hemric和J.M. Chalovich，1988年，《生物化学杂志》263卷，1878 - 1885页；L. Velaz、R.H. Ingraham和J.M. Chalovich，1990年，《生物化学杂志》265卷，2929 - 2934页）。根据这些数据，我们推测钙调蛋白是肌球蛋白亚片段-1与肌动蛋白结合的竞争性抑制剂。为了证实这一结果，我们现在展示了在肌动蛋白浓度增加时，钙调蛋白对（A1）S-1水解ATP的稳态参数的影响。采用低离子强度条件以最大化（A1）S-1与肌动蛋白之间的相互作用。在存在和不存在平滑肌原肌球蛋白的情况下，钙调蛋白均使kcat降低两倍，使KATPase变化超过12倍。因此，在存在和不存在原肌球蛋白的情况下，钙调蛋白与肌动蛋白结合的竞争都导致了ATP酶活性的降低。
