权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Analysis of Activation Mechanism of Calpain on the Basis of its Tertiary Structure

基于钙蛋白酶三级结构的激活机制分析

基本信息

批准号：
12308032
负责人：
SUZUKI Koichi
金额：
$ 21.7万
依托单位：
Tokyo Metropolitan Institute of Gerontology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12308032/
关键词：
Calpain Calcium Protease X ray structural analysis Higher order structure of protein Activation mechanism Phopsholipids 蛋白質の構造変化リン脂質

项目摘要

Calpain is composed of two subunits, 80K and 30K, but 80K itself expresses full enzyme activity. The results of X ray analysis of calpain in the absence of Ca, show that the protease domain (II) of 80K comprising domains (I-IV) is composed of two sub-domains, II and Iib, which are quite similar to those in other cysteine proteases. However, they are separated and the proper active-site that exists between the two sub-domains is not formed. The activation mechanism corresponding to how these two sub-domains come closer to forn proper active-site by conformational changes induced by Ca, was examined. Two sub-domains are separated mainly by two kinds of interactions, interaction between I and cahnodulin domain (VI) in 30K, and interaction of lib with III. Upon binding Ca to VI and III, these interactions are disrupted resulting in large conformational changes that make two sub-domains come closer. Further, minor conformational changes induced by Ca binding to II are essential to make II active. Ill plays very important roles in this activation ; acting as the novel Ca binding sites, and also as the phopspholipids binding domain that determines cellular localization of active calpain. A novel Ca binding site was also predicted in II but its precise location is still unknown. To know the structure of Ca-bound form of calpain is essential to understand the final activation mechanism. Further experiments are now in progress along this line.

钙蛋白酶由80 K和30 K两个亚基组成，但80 K本身表达完整的酶活性。无钙条件下钙蛋白酶的X射线分析结果表明，80 K蛋白酶结构域（II）由II和Iib两个亚结构域组成，这两个亚结构域与其它半胱氨酸蛋白酶十分相似。然而，它们是分开的，并且存在于两个子域之间的适当的活性位点没有形成。探讨了Ca诱导的构象变化使这两个亚结构域更接近合适的活性位点的激活机制。两个亚结构域主要通过两种相互作用分开，在30 K下I与钙调素结构域（VI）之间的相互作用，以及lib与III之间的相互作用。当Ca与VI和III结合时，这些相互作用被破坏，导致大的构象变化，使两个子域更接近。此外，轻微的构象变化引起的钙结合到II是必不可少的，使II活性。III在这种激活中起着非常重要的作用;作为新的Ca结合位点，也作为磷脂结合结构域，决定活性钙蛋白酶的细胞定位。一个新的钙结合位点也预测在II，但其精确的位置仍然是未知的。了解钙结合型钙蛋白酶的结构对于理解其最终的激活机制至关重要。沿着这条路线正在进行进一步的实验。