Sellers James R, Wang Fei, Chantler Peter D
Laboratory of Molecular Cardiology, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
J Muscle Res Cell Motil. 2003;24(8):579-85. doi: 10.1023/b:jure.0000009969.04562.58.
Trifluoperazine, a calmodulin antagonist, has recently been shown to inhibit the MgATPase activity of scallop myosin in the absence of light chain dissociation (Patel et al. (2000) J Biol Chem 275: 4880-4888). To investigate the generality of this observation and the mechanism by which it occurs, we have examined the ability of trifluoperazine to inhibit the enzymatic properties of other conventional and unconventional myosins. We show that trifluoperazine can inhibit the actin-activated MgATPase activity of rabbit skeletal muscle myosin II heavy meromyosin (HMM), phosphorylated turkey gizzard smooth muscle myosin II HMM, phosphorylated human nonmuscle myosin IIA HMM and myosin V subfragment-1 (S1). In all cases half maximal inhibition occurred at 50-75 microM trifluoperazine while light chains (myosin II) or calmodulin (myosin V) remained associated with the heavy chains. In vitro motility of all myosins tested was completely inhibited by trifluoperazine. Chymotryptic digestion of baculovirus-expressed myosin V HMM possessing only two calmodulin binding sites yielded a minimal motor fragment with no bound calmodulin. The MgATPase of this fragment was inhibited by trifluoperazine over the same range of concentrations as the S1 fragment of myosin.
三氟拉嗪是一种钙调蛋白拮抗剂，最近的研究表明，在轻链不解离的情况下，它能抑制扇贝肌球蛋白的MgATP酶活性（帕特尔等人，《生物化学杂志》，2000年，第275卷，第4880 - 4888页）。为了研究这一现象的普遍性及其发生机制，我们检测了三氟拉嗪抑制其他传统和非传统肌球蛋白酶活性的能力。我们发现，三氟拉嗪能够抑制兔骨骼肌肌球蛋白II重酶解肌球蛋白（HMM）、磷酸化火鸡肌胃平滑肌肌球蛋白II HMM、磷酸化人非肌肉肌球蛋白IIA HMM以及肌球蛋白V亚片段1（S1）的肌动蛋白激活的MgATP酶活性。在所有情况下，半最大抑制浓度为50 - 75微摩尔三氟拉嗪，此时轻链（肌球蛋白II）或钙调蛋白（肌球蛋白V）仍与重链结合。三氟拉嗪完全抑制了所有测试肌球蛋白的体外运动能力。对仅具有两个钙调蛋白结合位点的杆状病毒表达的肌球蛋白V HMM进行胰凝乳蛋白酶消化，产生了一个没有结合钙调蛋白的最小运动片段。该片段的MgATP酶在与肌球蛋白S1片段相同的浓度范围内受到三氟拉嗪的抑制。
