Recombinant motor domain constructs of Chara corallina myosin display fast motility and high ATPase activity
Recombinant motor domain constructs of Chara corallina myosin display fast motility and high ATPase activity
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DOI：
10.1016/j.bbrc.2003.10.202
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发表时间：
2003-12-26
影响因子：
3.1
通讯作者：
Yamamoto, K
中科院分区：
文献类型：
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作者：
Ito, K;Kashiyama, T;Yamamoto, K
珊瑚轮状肌球蛋白(CCM)快速运动的机制和结构特征尚未阐明。可提纯至均一的天然CCM产量低是造成这一现象的主要原因。在这里，我们描述了重组CCM运动域的表达，它支持肌动蛋白细丝在体外运动试验中的快速移动。没有轻链结合位点的CCM运动域以8.8um/S的速度30°C移动肌动蛋白细丝，带有由两个α-肌动蛋白重复序列组成的人工杠杆臂的CCM运动域以16.2um/S的速度移动肌动蛋白细丝。这两个结构域都显示出高的肌动蛋白激活的ATPase活性(类似于500pI/S/头)，这表明水解步骤非常快。我们的结果提供了一个很好的系统来剖析特定的结构和功能特征，以区分负责快速胞浆流动的肌球蛋白。(C)2003 Elsevier Inc.保留所有权利。
The mechanism and structural features that are responsible for the fast motility of Chara corallina myosin (CCM) have not been elucidated, so far. The low yields of native CCM that can be purified to homogeneity were the major reason for this. Here, we describe the expression of recombinant CCM motor domains, which support the fast movement of actin filaments in an in vitro motility assay. A CCM motor domain without light chain binding site moved actin filaments at a velocity of 8.8 mum/s at 30 degreesC and a CCM motor domain with an artificial lever arm consisting of two alpha-actinin repeats moved actin filaments at 16.2 mum/s. Both constructs displayed high actin-activated ATPase activities (similar to500 Pi/s/head), which is indicative of a very fast hydrolysis step. Our results provide an excellent system to dissect the specific structural and functional features that distinguish the myosin responsible for fast cytoplasmic streaming. (C) 2003 Elsevier Inc. All rights reserved.