Hemric M E, Chalovich J M
Department of Biochemistry, East Carolina University School of Medicine, Greenville, North Carolina 27858.
J Biol Chem. 1988 Feb 5;263(4):1878-85.
We have previously shown that inhibition of the ATPase activity of skeletal muscle myosin subfragment 1 (S1) by caldesmon is correlated with the inhibition of S1 binding in the presence of ATP or pyrophosphate (Chalovich, J., Cornelius, P., and Benson, C. (1987) J. Biol Chem. 262, 5711-5716). In contrast, Lash et al. (Lash, J., Sellers, J., and Hathaway, D. (1986) J. Biol. Chem. 261, 16155-16160) have shown that the inhibition of ATPase activity of smooth muscle heavy meromyosin (HMM) by caldesmon is correlated with an increase in the binding of HMM to actin in the presence of ATP. We now show, in agreement, that caldesmon does increase the binding of smooth muscle HMM to actin-tropomyosin while decreasing the ATPase activity. The effect of caldesmon on the binding of smooth HMM is reversed by Ca2+-calmodulin. Caldesmon strengthens the binding of smooth S1.ATP and skeletal HMM.ATP to actin-tropomyosin but to a lesser extent than smooth HMM.ATP. Furthermore, this increase in binding of smooth S1.ATP and skeletal HMM.ATP does not parallel the inhibition of ATPase activity. In contrast, in the absence of ATP, all smooth and skeletal myosin subfragments compete with caldesmon for binding to actin. Thus, the effect that caldesmon has on the binding of myosin subfragments to actin-tropomyosin depends on the source of myosin, the type of subfragment, and the nucleotide present. The inhibition of actin-activated ATP hydrolysis by caldesmon, however, is not greatly different for different smooth and skeletal myosin subfragments. Evidence is presented that caldesmon inhibits actin-activated ATP hydrolysis by attenuating the productive interaction between myosin and actin that normally accelerates ATP hydrolysis. The increased binding seen by some myosin subfragments, in the presence of ATP, may be due to binding of these subfragments to a nonproductive site on actin-caldesmon. The subfragments which show an increase in binding in the presence of ATP and caldesmon appear to bind directly to caldesmon as demonstrated by affinity chromatography.
我们之前已经表明，钙调蛋白对骨骼肌肌球蛋白亚片段1（S1）ATP酶活性的抑制与在ATP或焦磷酸存在下S1结合的抑制相关（查洛维奇，J.，科尼利厄斯，P.，和本森，C.（1987）《生物化学杂志》262，5711 - 5716）。相比之下，拉什等人（拉什，J.，塞勒斯，J.，和哈撒韦，D.（1986）《生物化学杂志》261，16155 - 16160）已经表明，钙调蛋白对平滑肌重酶解肌球蛋白（HMM）ATP酶活性的抑制与在ATP存在下HMM与肌动蛋白结合的增加相关。我们现在一致表明，钙调蛋白确实增加了平滑肌HMM与肌动蛋白 - 原肌球蛋白的结合，同时降低了ATP酶活性。钙调蛋白对平滑肌HMM结合的影响可被Ca2 + - 钙调蛋白逆转。钙调蛋白增强了平滑肌S1.ATP和骨骼肌HMM.ATP与肌动蛋白 - 原肌球蛋白的结合，但程度小于平滑肌HMM.ATP。此外，平滑肌S1.ATP和骨骼肌HMM.ATP结合的这种增加与ATP酶活性的抑制并不平行。相比之下，在没有ATP的情况下，所有平滑肌和骨骼肌肌球蛋白亚片段都与钙调蛋白竞争与肌动蛋白的结合。因此，钙调蛋白对肌球蛋白亚片段与肌动蛋白 - 原肌球蛋白结合的影响取决于肌球蛋白的来源、亚片段的类型以及存在的核苷酸。然而，钙调蛋白对不同平滑肌和骨骼肌肌球蛋白亚片段肌动蛋白激活的ATP水解的抑制作用并没有太大差异。有证据表明，钙调蛋白通过减弱肌球蛋白和肌动蛋白之间通常加速ATP水解的有效相互作用来抑制肌动蛋白激活的ATP水解。在ATP存在下，一些肌球蛋白亚片段观察到的结合增加可能是由于这些亚片段与肌动蛋白 - 钙调蛋白上的非生产性位点结合。如亲和色谱所示，在ATP和钙调蛋白存在下显示结合增加的亚片段似乎直接与钙调蛋白结合。
