Mouse Myosin-19 Is a Plus-end-directed, High-duty Ratio Molecular Motor
Mouse Myosin-19 Is a Plus-end-directed, High-duty Ratio Molecular Motor
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Mouse Myosin-19 是一种正端定向、高占空比分子马达
DOI：
10.1074/jbc.m114.569087
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发表时间：
2014-06-27
影响因子：
4.8
通讯作者：
Li, Xiang-dong
中科院分区：
文献类型：
--
作者：
Lu, Zekuan;Ma, Xiao-Nan;Li, Xiang-dong
XIX类肌球蛋白（Myo 19）是脊椎动物特有的非常规肌球蛋白，负责线粒体的运输。为了表征Myo 19的生化特性，我们使用杆状病毒/Sf 9表达系统制备了含有马达结构域和IQ基序的重组小鼠Myo 19截短构建体。我们确定平滑肌/非肌肉肌球蛋白-2的调节轻链（RLC）为Myo 19的轻链。肌动蛋白激活的ATP酶活性和肌动蛋白滑行速度的Myo 19截短的结构是约三分之一和六分之一的肌球蛋白-5a，分别。Myo 19对肌动蛋白的表观亲和力与肌球蛋白-5a的表观亲和力大致相同。肌球蛋白轻链激酶可以磷酸化结合Myo 19的RLC，但这种磷酸化对肌动蛋白激活的ATP酶活性和肌动蛋白滑行活性的Myo 19截短的结构几乎没有影响。使用双荧光标记的肌动蛋白丝，我们确定Myo 19是一个正末端定向分子马达。我们发现，类似的高占空比肌球蛋白，如肌球蛋白-5a，ADP释放速率与最大肌动蛋白激活的ATP酶活性的Myo 19，表明ADP释放是一个ratelimiting步骤的ATP酶循环的acto-Myo 19。ADP强烈抑制肌动蛋白激活的ATP酶活性和肌动蛋白滑行活性的Myo 19-截断的结构。基于上述结果，我们得出结论，Myo 19是一个高占空比的分子马达运动到肌动蛋白丝的正端。
Class XIX myosin (Myo19) is a vertebrate-specific unconventional myosin, responsible for the transport of mitochondria. To characterize biochemical properties of Myo19, we prepared recombinant mouse Myo19-truncated constructs containing the motor domain and the IQ motifs using the baculovirus/Sf9 expression system. We identified regulatory light chain (RLC) of smooth muscle/non-muscle myosin-2 as the light chain of Myo19. The actin-activated ATPase activity and the actin-gliding velocity of Myo19-truncated constructs were about onethird and one-sixth as those of myosin-5a, respectively. The apparent affinity of Myo19 to actin was about the same as that of myosin-5a. The RLCs bound to Myo19 could be phosphorylated by myosin light chain kinase, but this phosphorylation had little effect on the actin-activated ATPase activity and the actin-gliding activity of Myo19-truncated constructs. Using dual fluorescence-labeled actin filaments, we determined that Myo19 is a plus-end-directed molecular motor. We found that, similar to that of the high-duty ratio myosin, such as myosin-5a, ADP release rate was comparable with the maximal actin-activated ATPase activity of Myo19, indicating that ADP release is a ratelimiting step for the ATPase cycle of acto-Myo19. ADP strongly inhibited the actin-activated ATPase activity and actin-gliding activity of Myo19-truncated constructs. Based on the above results, we concluded that Myo19 is a high-duty ratio molecular motor moving to the plus-end of the actin filament.