Folding and Activation of Client Proteins by the Hsp90 Molecular Chaperone Machine

分子伴侣调节细胞蛋白质从合成到正确折叠或降解的折叠过程。大多数分子伴侣帮助折叠大部分未展开的蛋白质。丰富的、必需的分子伴侣蛋白Hsp90（热休克蛋白，90 kDa）在伴侣蛋白中是独一无二的，因为它优先与几乎完全折叠的蛋白质相互作用，并且是蛋白质活性所必需的。需要Hsp90才能发挥功能的客户蛋白很少或没有序列或结构同源性，并且对于Hsp90如何促进一组不同细胞客户蛋白的激活知之甚少。该项目的目标是确定Hsp90与两种非常不同的客户蛋白之间相互作用的位点。一组Hsp90的突变形式将被用作识别和描述Hsp90与蛋白激酶和转录因子相互作用所需的关键特征的工具。一旦在体内发现破坏与两种不同的客户蛋白相互作用的突变，进一步的研究将确定这些突变是否会利用细胞裂解物和纯化组分破坏hsp90 -客户蛋白复合物的体外组装。进一步的研究将集中于确定选择的Hsp90突变是否通过阻止Hsp90与调控共伴侣蛋白的相互作用间接破坏客户蛋白的相互作用。该研究项目将有助于澄清Hsp90和客户蛋白之间的接触位点，并进一步了解Hsp90和co-chaperones如何合作促进多种客户蛋白的激活。该项目将对了解Hsp90如何与多种客户蛋白相互作用具有广泛的影响。该项目每年将允许一名研究生和两名本科生进行独立的研究项目。所有学生都将有机会在实验室会议、本科生研究海报论坛和/或其他科学会议中发展组织和展示数据的技能。本实验室以外的研究生和本科生将通过微生物学、分子生物学和生物化学系以及生殖生物学中心（与华盛顿州立大学的联合项目）的教员之间的科学交流和研讨会接触到这项工作。通过NIH-INBRE项目和作为多州WWAMI医学院项目一部分的UDOC项目，来自代表性不足和少数群体的更多本科生和高中生也将接触到实验室的研究。

Molecular chaperones regulate the folding of cellular proteins from synthesis until they are properly folded or degraded. The majority of molecular chaperones assist the folding of proteins that are mostly unfolded. The abundant, essential molecular chaperone Hsp90 (heat shock protein, 90 kDa) is unique among chaperones because it preferentially interacts with and is required for the activity of proteins that are almost completely folded. The client proteins that require Hsp90 for function have little or no sequence or structural homology, and little is known about how Hsp90 promotes activation of a select group of diverse cellular client proteins. The goal of this project is to identify the sites of interaction between Hsp90 and two very different client proteins. A panel of mutant forms of Hsp90 will be used as tools to identify and describe critical features of Hsp90 required for interaction with a protein kinase and a transcription factor. Once mutations that disrupt the interaction with two distinct client proteins in vivo are identified, further research will determine whether the mutations disrupt the in vitro assembly of Hsp90-client protein complexes using cell lysates and purified components. Additional research will focus on establishing whether select Hsp90 mutations indirectly disrupt client protein interaction by preventing interaction of Hsp90 with regulatory co-chaperone proteins. This research project will lead to clarification of the contact sites between Hsp90 and client proteins and a greater understanding of how Hsp90 and co-chaperones cooperate to promote activation of a diverse group of client proteins. This project will have broad impacts on understanding how Hsp90 interacts with diverse client proteins. This project will enable one graduate student and two undergraduate students per year to conduct independent research projects. All students will have the opportunity to develop skills in organizing and presenting data during laboratory meetings, undergraduate research poster forums and/or additional scientific conferences. Additional graduate and undergraduate students outside of this lab will be exposed to this work through scientific exchanges and seminars among faculty members within the Microbiology, Molecular Biology and Biochemistry department, and the Center for Reproductive Biology, which is a joint program with Washington State University. Through the NIH-INBRE program and the UDOC program that is part of the multi-state WWAMI medical school program, additional undergraduate students and high school students from underrepresented and minority groups will also be exposed to research in the laboratory.