Kinetic signatures of myosin-5B, the motor involved in microvillus inclusion disease
Kinetic signatures of myosin-5B, the motor involved in microvillus inclusion disease
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DOI：
10.1074/jbc.m117.801456
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发表时间：
2017-11-03
影响因子：
4.8
通讯作者：
Sellers, James R.
中科院分区：
文献类型：
--
作者：
Heissler, Sarah M.;Chinthalapudi, Krishna;Sellers, James R.
肌球蛋白-5B是一种普遍存在的分子马达，在细胞内循环途径中运输内膜系统的货物囊泡。肌球蛋白-5B功能障碍导致先天性肠病微绒毛包涵体病，强调其在细胞内稳态中的重要性。在这里，我们描述了肌球蛋白-5B与F-肌动蛋白，核苷酸和吡唑并嘧啶化合物myoVin-1的相互作用。我们发现，单头肌球蛋白-5B是一个中间占空比电机与动力ATP酶循环，是由磷酸盐的释放速率限制。第二个头部的存在会在肌球蛋白-5B二聚体中产生应变和门控，通过降低肌动蛋白激活的ADP释放速率来改变动力学特征，从而成为限速。这种动力学转变为高占空比电机是肌球蛋白-5B在细胞再循环途径中的拟议运输功能的先决条件。此外，我们表明，小分子化合物myoVin-1在体外抑制肌球蛋白-5B的酶活性和功能活性。肌动蛋白激活的稳态ATP酶活性和滑动速度的部分抑制表明，在探测myoVin-1对细胞中肌球蛋白-5依赖性转运过程的影响时，应谨慎使用。
Myosin-5B is a ubiquitous molecular motor that transports cargo vesicles of the endomembrane system in intracellular recycling pathways. Myosin-5B malfunction causes the congenital enteropathy microvillus inclusion disease, underlining its importance in cellular homeostasis. Here we describe the interaction of myosin-5B with F-actin, nucleotides, and the pyrazolopyrimidine compound myoVin-1. We show that single-headed myosin-5B is an intermediate duty ratio motor with a kinetic ATPase cycle that is rate-limited by the release of phosphate. The presence of a second head generates strain and gating in the myosin-5B dimer that alters the kinetic signature by reducing the actin-activated ADP release rate to become rate-limiting. This kinetic transition into a high-duty ratio motor is a prerequisite for the proposed transport function of myosin-5B in cellular recycling pathways. Moreover, we show that the small molecule compound myoVin-1 inhibits the enzymatic and functional activity of myosin-5B in vitro. Partial inhibition of the actin-activated steady-state ATPase activity and sliding velocity suggests that caution should be used when probing the effect of myoVin-1 on myosin-5-dependent transport processes in cells.