Myosin analyses using recombinant motor domain constructs of Chara coralline myosin

使用 Chara 珊瑚肌球蛋白的重组运动结构域结构进行肌球蛋白分析

• 批准号: 15570133
• 负责人: ITO Kohji
• 金额: $ 1.98万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15570133/
• 关键词: mosin; motor; Characean; Cytoplasmic streaming; ATPas activity; in vitro motility assay; actin; in vitro motility assay; 分子モーター

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 μm/s at 30℃ and a CCM motor domain with an artificial lever arm consisting of two alpha-actinin repeats moved actin filaments at 16.2 μm/s. Both constructs displayed high actin-activated ATPase activities (〜500 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.

迄今为止，导致轮藻肌球蛋白（CCM）快速运动的机制和结构特征尚未阐明。可以纯化至均一的天然CCM的低产量是其主要原因。在这里，我们描述了重组CCM马达结构域的表达，它支持肌动蛋白丝在体外运动试验中的快速运动。无轻链结合位点的CCM马达结构域在30℃下以8.8 μm/s的速度移动肌动蛋白丝，而有两个α-辅肌动蛋白重复序列组成的人工杠杆臂的CCM马达结构域以16.2 μm/s的速度移动肌动蛋白丝。两种构建体均显示出高肌动蛋白激活的ATP酶活性（约500 Pi/s/头），这表明水解步骤非常快。我们的研究结果提供了一个很好的系统，解剖的具体结构和功能的特点，区分负责快速细胞质流的肌球蛋白。

项目成果

Roles of hydrophobic triplet in the motor domain of myosin in the interaction between myosin and action.

肌球蛋白运动结构域中疏水三联体在肌球蛋白与动作相互作用中的作用。

• 发表时间: 2003
• 期刊: J.Biochem. 134
• 作者: Hachikubo Y.;Ito K.;Keiichi Yamamoto
• 通讯作者: Keiichi Yamamoto

Kohji Ito, Taro Q.P.Uyeda, Yoshikazu Suzuki, Kazuo Sutoh, Keiichi Yamamoto: "Requirement of domain-domain interaction for conformational change and functional ATP hydrolysis in myosin."J.Biol.Chem.. 278・33. 31049-31057 (2003)

Kohji Ito、Taro Q.P.Uyeda、Yoshikazu Suzuki、Kazuo Sutoh、Keiichi Yamamoto：“肌球蛋白构象变化和功能性 ATP 水解的结构域相互作用的要求。J.Biol.Chem.. 278・31057”（2003 年） ）

Recombinant motor domain constructs of Chara corallina myosin display fast motility and high ATPase activity

• DOI: 10.1016/j.bbrc.2003.10.202
• 发表时间: 2003-12-26
• 期刊: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
• 影响因子: 3.1
• 作者: Ito, K;Kashiyama, T;Yamamoto, K
• 通讯作者: Yamamoto, K

Susceptibility of chara myosin to SH regents.

轮藻肌球蛋白对 SH 试剂的敏感性。

• 发表时间: 2003
• 期刊: Plant Cell Physiol. 44
• 作者: Seki M.;Awata J.;Shimada K.;Kashiyama T;Ito K.;Yamamoto K.
• 通讯作者: Yamamoto K.

The motility of Chara corallina myosin was inhibited reversibly by 2,3-butanedione monoxime (BDM)

• DOI: 10.1093/pcp/pch154
• 发表时间: 2004-09-01
• 期刊: PLANT AND CELL PHYSIOLOGY
• 影响因子: 4.9
• 作者: Funaki, K;Nagata, A;Yamamoto, K
• 通讯作者: Yamamoto, K