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

• 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
• 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

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.

热休克蛋白90是真核生物中一种重要的分子伴侣，在机体的活性产生和维持中起重要作用 选择一组与信号过程不成比例地联系的基质或客户端 包括许多促进癌症和真菌抗药性出现的基因。为这些 由于这些原因，操纵Hsp90分子伴侣系统的抑制剂策略有可能带来许多潜在的 治疗益处。靶向Hsp90的ATP酶位点的抑制剂有效地限制了Hsp90的活性。 真菌耐药性的出现，并显示出作为抗癌剂的前景。 然而，ATP酶抑制剂阻断了Hsp90的所有已知功能，导致不良副作用。 方面的影响.在我们之前的工作中，我们在2015年创建了一个包含所有可能突变的综合文库， Hsp90的709个位点中的每一个，并量化每个变体对酵母生长的影响 速率作为所有客户蛋白质上集成的整体网络功能的读数。我们将建立在 本研究旨在探讨热休克蛋白90与客户端的相互作用机制。我们将确定四个具体的 客户受到Hsp90突变的影响，这将确定Hsp90上可能发生突变的位点。 针对特定客户的攻击我们将对特定客户端进行深度突变扫描 蛋白质来鉴定决定Hsp90依赖性的生物物理特征。另外我们 已经开发了一种新的方法来量化Hsp90突变的显性效应， 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了重要的机制见解， 在生物学和医学上都很重要