A Dictyostelium myosin II lacking a proximal 58-kDa portion of the tail is functional in vitro and in vivo.
A Dictyostelium myosin II lacking a proximal 58-kDa portion of the tail is functional in vitro and in vivo.
复制标题
缺少尾部近端 58 kDa 部分的盘基网柄菌肌球蛋白 II 在体外和体内均具有功能。
DOI：
10.1091/mbc.3.12.1455
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发表时间：
1992
影响因子：
3.3
通讯作者：
Spudich,JA
中科院分区：
文献类型：
--
作者：
Kubalek,EW;Uyeda,TQ;Spudich,JA
我们使用分子遗传学方法删除了521个氨基酸残基的近端部分的网囊藻肌球蛋白II的尾巴。缺失包括大约40%的尾部，包括S2-LMM连接，这是一个在肌肉肌球蛋白II中被认为对收缩很重要的区域。在我们的体外试验中，突变型肌球蛋白II的功能与野生型肌球蛋白II没有区别。在缺乏ATP的情况下，它以典型的僵硬构型与肌动蛋白结合，并以正常的盐依赖性方式形成细丝。在体外运动性测定中，突变体肌球蛋白II的单体和丝状形式在30摄氏度下以2.4微米/秒的速度移位肌动蛋白丝，与野生型肌球蛋白II相似。突变型肌球蛋白II在体内也有功能。细胞表达的突变体肌球蛋白II的天然肌球蛋白II的地方执行肌球蛋白II依赖的活动，如胞质分裂和子实体的形成，虽然效率低下。突变体细胞在悬浮液中的生长产生许多大的多核细胞，表明胞质分裂常常失败。大部分子实体的形态也是异常的。这些结果表明，该地区的肌球蛋白II尾巴是不需要的运动活动，但它的存在是必要的最佳功能在体内。
We used molecular genetic approaches to delete 521 amino acid residues from the proximal portion of the Dictyostelium myosin II tail. The deletion encompasses approximately 40% of the tail, including the S2-LMM junction, a region that in muscle myosin II has been proposed to be important for contraction. The functions of the mutant myosin II are indistinguishable from the wild-type myosin II in our in vitro assays. It binds to actin in a typical rigor configuration in the absence of ATP and it forms filaments in a normal salt-dependent manner. In an in vitro motility assay, both monomeric and filamentous forms of the mutant myosin II translocate actin filaments at 2.4 microns/s at 30 degrees C, similar to that of wild-type myosin II. The mutant myosin II is also functional in vivo. Cells expressing the mutant myosin II in place of the native myosin II perform myosin II-dependent activities such as cytokinesis and formation of fruiting bodies, albeit inefficiently. Growth of the mutant cells in suspension gives rise to many large multinucleated cells, demonstrating that cytokinesis often fails. The majority of the fruiting bodies are also morphologically abnormal. These results demonstrate that this region of the myosin II tail is not required for motile activities but its presence is necessary for optimum function in vivo.