Structural Studies of Clp/Hsp 100 Molecular Chaperones

描述（由申请方提供）：蛋白质必须正确折叠以获得生物学功能。同时，蛋白质聚集和错误折叠是许多毁灭性人类疾病的关键因素，例如阿尔茨海默病、朊病毒介导的感染、II型糖尿病和囊性纤维化。虽然“常规”分子伴侣通过促进“正向”折叠或防止蛋白质聚集来辅助蛋白质折叠，但它们不能促进已经聚集的蛋白质如淀粉样蛋白的解聚，淀粉样蛋白与某些人类疾病相关。 细菌ClpB及其真核同源物Hsp 104是热休克反应的必需蛋白，形成大的环状结构，并且属于与多种细胞活性（AAA+）相关的ATP酶的Hsp 100家族。与任何其他分子伴侣（包括Hsp 100家族的其他成员）不同，ClpB/Hsp 104具有促进已经聚集的应激损伤蛋白解聚的显著能力。由于缺乏高分辨率的结构信息，其潜在机制目前尚不清楚。 长期目标是了解ClpB/Hsp 104家族成员促进应激损伤蛋白解聚的分子机制。本研究的目标将通过以下具体目标来实现：（1）使用X射线晶体学来解决CIpB/Hsp 104的高分辨率晶体结构，（2）通过冷冻电子显微镜来阐明0.6-MDa CIpB/Hsp 104组装体的三维结构，以及（3）确定CIpB/Hsp 104识别和结合模型底物的结构基础。这些研究将通过突变和生化实验来补充，以测试从这些结构推断的假设。这些方法的结合将提供一个详细的机制的理解CIpB/Hsp 104的结构-功能关系，并可能激发新技术的设计，可能导致一个潜在的治疗人类朊病毒和淀粉样蛋白疾病。

DESCRIPTION (provided by applicant): Proteins must fold correctly in order to attain biological function. Concurrently, protein aggregation and misfolding are key contributors to many devastating human diseases such as Alzheimer's disease, prion-mediated infections, type II diabetes, and cystic fibrosis. While "conventional" molecular chaperones assist protein folding by promoting the "forward" folding or preventing protein aggregation, they are unable to promote the disaggregation of already aggregated proteins such as amyloids, which are associated with certain human diseases. Bacterial CIpB and its eukaryotic homolog Hsp104 are essential proteins of the heat-shock response, form large ring-like structures, and belong to the Hsp100 family of ATPases associated with diverse cellular activities (AAA+). Unlike any other chaperone, including other members of the Hsp100 family, CIpB/Hsp104 has the remarkable ability to promote the disaggregation of already aggregated, stress-damaged proteins. The underlying mechanism is currently unknown due to the lack of high-resolution structural information. The long-term objective is to understand the molecular mechanism by which members of the CIpB/Hsp104 family promote the disaggregation of stress-damaged proteins. The goals of this research will be pursued through the following specific aims: (1) to solve the high-resolution crystal structure of CIpB/Hsp104 using X-ray crystallography, (2) to elucidate the three-dimensional structure of the 0.6-MDa CIpB/Hsp104 assembly by cryo-electron microscopy, and (3) to determine the structural basis by which CIpB/Hsp104 recognizes and binds model substrates. These studies will be complemented by mutational and biochemical experiments to test the hypotheses inferred from these structures. The combination of these approaches will provide a detailed mechanistic understanding of the structure-function relationship of CIpB/Hsp104, and may inspire the design of novel technology that could lead to a potential cure for human prion and amyloid diseases.