...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Molecular biology of the cell >Interaction of dynamin with microtubules: its structure and GTPase activity investigated by using highly purified dynamin.
【24h】

Interaction of dynamin with microtubules: its structure and GTPase activity investigated by using highly purified dynamin.

机译：动力蛋白与微管的相互作用：使用高度纯化的动力蛋白研究其结构和GTPase活性。

获取原文
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

We purified a large amount of dynamin with high enzymatical activity from rat brain tissue by a new procedure. Dynamin 0.48 mg was obtained from 20 g of rat brain. The purity of dynamin was almost 98%. Dynamin plays a role of GTPase rather than ATPase. In the absence of microtubules, Michaelis constant (Km) and maximum velocity (Vmax) for dynamin GTPase were 370 microM and 0.25 min-1, respectively, and in their presence, both were significantly accelerated up to 25 microM and 5.5 min-1. On the other hand, the ATPase activity was very low in the absence of microtubules, and even in their presence, Km and Vmax for dynamin ATPase were 0.2 mM and 0.91 min-1. Despite slow GTPase turnover rate in the absence of microtubules, binding of GTP and its nonhydrolizing analogues was very fast, indicating that GTP binding step is not rate limiting. Dynamin did not cause a one-directional consistent microtubule sliding movement just like kinesin or dynein in the presence of 2 mM ATP or 2 mM GTP. We observed the molecular structure of dynamin with low-angle rotary shadowing technique and revealed that the dynamin molecule is globular in shape. Gel filtration assay revealed that these globules were the oligomers of 100-kDa dynamin polypeptide. Dynamin bound to microtubules with a 1:1 approximately 1.2 molar ratio in the absence of GTP. Quick-freeze deep-etch electron microscopy of the dynamin-microtubule complex showed that dynamin decorates the surface of microtubules helically, like a screw bolt, very orderly and tightly with 11.4 +/- 0.9 (SD)nm period. Contrary to the previous report, microtubules make bundles by the attachment of the dynamin helixes around each adjacent microtubule, and no cross-bridge formation was observed.
机译：我们通过一种新方法从大鼠脑组织中纯化了大量具有高酶促活性的动力蛋白。从20克大鼠大脑中获得了0.48毫克的动态发电机。 dynamin的纯度几乎为98％。动力蛋白起着GTPase而不是ATPase的作用。在没有微管的情况下，动态GTPase的米氏常数（Km）和最大速度（Vmax）分别为370 microM和0.25 min-1，并且在存在它们的情况下，两者均被显着加速至25 microM和5.5 min-1。另一方面，在没有微管的情况下，ATPase的活性非常低，即使在存在微管的情况下，dynamin ATPase的Km和Vmax也为0.2 mM和0.91 min-1。尽管在不存在微管的情况下GTP酶的转换速率较慢，但GTP及其非水解类似物的结合非常快，这表明GTP结合步骤不受速率限制。在存在2 mM ATP或2 mM GTP的情况下，动力蛋白不会像驱动蛋白或动力蛋白那样引起单向一致的微管滑动运动。我们用低角度旋转遮蔽技术观察了动力蛋白的分子结构，发现该动力蛋白分子呈球形。凝胶过滤试验表明这些小球是100-kDa dynamin多肽的寡聚体。在没有GTP的情况下，动力蛋白以1：1约1.2的摩尔比结合微管。快速冷冻的电镜-微管复合物的深蚀刻电子显微镜显示，电镜的螺旋状装饰微管表面（如螺栓）非常有序且紧密，周期为11.4 +/- 0.9（SD）nm。与先前的报告相反，微管通过围绕每个相邻微管的动力螺旋的附着而形成束，并且未观察到过桥形成。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号