Ikebe M, Inagaki M, Naka M, Hidaka H
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
J Biol Chem. 1988 Aug 5;263(22):10698-704.
The rate of phosphorylation and dephosphorylation of smooth muscle myosin by myosin light chain kinase and by two myosin light chain phosphatases (gizzard phosphatase IV and aorta phosphatase) are measured in various conditions; the relationship between the rate of phosphorylation and dephosphorylation of myosin and the myosin conformation is also studied. The rate of dephosphorylation of myosin was completely inhibited in the presence of 1 mM MgCl2 and ATP at low ionic strength where phosphorylated myosin forms a folded conformation. The inhibition was released when myosin formed either an extended monomer or filaments. The rate of phosphorylation of myosin was also affected by the conformation of myosin. The rate for a folded myosin was slower than those for an extended monomer and filamentous myosin. The phosphorylation and dephosphorylation of heavy meromyosin, subfragment-1, and the isolated 20,000-dalton light chain are not inhibited at low ionic strength, and the rate of phosphorylation and dephosphorylation was decreased with increasing ionic strength. KCl dependence of the rate of phosphorylation and dephosphorylation of myosin was normalized by using KCl dependence of subfragment-1, and it was found that the marked inhibition of the rate of phosphorylation and dephosphorylation of myosin is closely related to the change from an extended to a folded conformation of myosin.
在不同条件下测量了肌球蛋白轻链激酶以及两种肌球蛋白轻链磷酸酶（砂囊磷酸酶IV和主动脉磷酸酶）对平滑肌肌球蛋白的磷酸化和去磷酸化速率；还研究了肌球蛋白磷酸化和去磷酸化速率与肌球蛋白构象之间的关系。在低离子强度下，当磷酸化肌球蛋白形成折叠构象时，1 mM MgCl2和ATP存在时肌球蛋白的去磷酸化速率被完全抑制。当肌球蛋白形成伸展的单体或细丝时，抑制作用解除。肌球蛋白的磷酸化速率也受其构象影响。折叠态肌球蛋白的磷酸化速率比伸展单体和丝状肌球蛋白的磷酸化速率慢。重酶解肌球蛋白、亚片段-1和分离出的20,000道尔顿轻链的磷酸化和去磷酸化在低离子强度下不受抑制，且磷酸化和去磷酸化速率随离子强度增加而降低。通过使用亚片段-1的氯化钾依赖性对肌球蛋白磷酸化和去磷酸化速率的氯化钾依赖性进行归一化处理，发现肌球蛋白磷酸化和去磷酸化速率的显著抑制与肌球蛋白从伸展构象转变为折叠构象密切相关。
