Relating protein interaction networks to physiology by systematic mutant analyses

通过系统突变分析将蛋白质相互作用网络与生理学联系起来

基本信息

批准号：
10224929
负责人：
DANIEL N BOLON
金额：
$ 35.18万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-02-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10224929
关键词：
ATP phosphohydrolase Antineoplastic Agents Biochemical Biological Biophysics Calcineurin Catalytic Domain Cells Client Complex Dependence Development Disease Dominant-Negative Mutation Drug Targeting Drug resistance Engineering Fungal Drug Resistance Genes Glucocorticoid Receptor Grant Growth Health Human Individual Libraries Link Malignant Neoplasms Maps Medical Molecular Chaperones Mutation Mycoses Outcome Phosphotransferases Physiology Point Mutation Positioning Attribute Process Proteins Publishing Reporter Resolution Role Route Sampling Signal Transduction Signaling Protein Site Sodium Chloride Structure System Technology Therapeutic Variant Work Yeasts base biological adaptation to stress calcineurin phosphatase combat fitness fungus human disease improved inhibitor/antagonist insight mutant mutation screening next generation sequencing novel strategies pressure promoter protein protein interaction side effect tool transcription factor

项目摘要

Hsp90 is an essential eukaryotic chaperone that helps to produce and maintain the active state of a select set of substrates or clients that are disproportionately linked to signaling processes including many that promote cancer and the emergence of drug resistance in fungi. For these reasons, inhibitor strategies to manipulate the Hsp90 chaperone system promise many potential therapeutic benefits. Inhibitors targeting the ATPase site of Hsp90 effectively limit the emergence of drug resistance in fungi and show promise as anti-cancer agents. However, ATPase inhibitors block all known functions of Hsp90, leading to undesirable side effects. In our previous work, we generated a comprehensive library of all possible mutations at each of the 709 positions in Hsp90 and quantified the effects of each variant on yeast growth rate as a readout of overall network function integrated over all client proteins. We will build on this work to investigate Hsp90-client interaction mechanism. We will determine how four specific clients are impacted by Hsp90 mutations, which will identify sites on Hsp90 that could be targeted to inhibit specific clients. We will perform deep mutational scanning on specific client proteins to identify the biophysical features that determine Hsp90 dependence. In addition, we have developed a novel approach to quantify the dominant effects of Hsp90 mutations, which we are using as a powerful new route to understand its mechanism of action. Our studies will provide important mechanistic insights into a protein-protein interaction network that is biologically and medically important.

Hsp90 是一种重要的真核伴侣，有助于产生和维持活性状态与信号传导过程不成比例地相关的一组选定的底物或客户包括许多促进癌症和真菌耐药性出现的因素。对于这些原因，操纵 Hsp90 伴侣系统的抑制剂策略有望带来许多潜力治疗益处。针对 Hsp90 ATP 酶位点的抑制剂可有效限制真菌中出现耐药性并显示出作为抗癌药物的前景。然而，ATPase 抑制剂会阻断 Hsp90 的所有已知功能，从而导致不良副作用影响。在我们之前的工作中，我们生成了一个包含所有可能突变的综合库对 Hsp90 中的 709 个位置中的每一个进行分析，并量化每个变体对酵母生长的影响速率作为所有客户端蛋白质集成的整体网络功能的读数。我们将在此基础上这项工作旨在研究 Hsp90-客户端交互机制。我们将确定四项具体措施客户受到 Hsp90 突变的影响，这将识别 Hsp90 上可能的位点有针对性地抑制特定客户。我们将对特定客户进行深度突变扫描蛋白质来识别决定 Hsp90 依赖性的生物物理特征。此外，我们开发了一种新方法来量化 Hsp90 突变的显着影响，我们正在使用一种强大的新途径来了解其作用机制。我们的研究将为蛋白质-蛋白质相互作用网络提供重要的机制见解具有重要的生物学和医学意义。