Molecular mechanism of the interaction of Hsp90 with cochaperones and clients

依赖于ATP的90 kDa热休克蛋白(Hsp90)是进化上高度保守的分子伴侣，在所有真核细胞中都是必不可少的。Hsp90通过与Hsp70分子伴侣和30多个辅分子伴侣蛋白的合作，控制着许多折叠蛋白的稳定性和活性，这些蛋白被称为客户，包括蛋白激酶、受体、转录因子和一系列不同的其他蛋白。许多Hsp90客户端是基本信号转导通路和调节网络的组成部分，当与Hsp90处于复杂状态或被Hsp90正确折叠时，它们只对上游信号做出反应。本项目的目的是通过(1)对Hsp90-辅助伴侣复合体和Hsp90-辅助伴侣-客户复合体的结构和功能进行分析，(2)筛选新的辅助伴侣和Hsp90系统的修饰物，以在酵母中成熟激酶，(3)分析翻译后修饰对人类Hsp90构象动力学和功能的影响，从而加深我们对Hsp90复杂机制的理解。在我们的研究中，我们将使用广泛的生化和生物物理方法，包括荧光技术、氢交换质谱仪(HX-MS)和蛋白质-蛋白质相互作用技术，以及面包酵母和哺乳动物细胞培养中的遗传学和分子生物学方法。

The ATP dependent 90 kDa heat shock proteins (Hsp90s) are evolutionarily highly conserved and very abundant molecular chaperone that are essential in all eukaryotic cells. In cooperation with Hsp70 chaperones and, depending on the organism, more than 30 cochaperones, Hsp90s control stability and activity of many folded protein, called clients, including protein kinases, receptors, transcription factors, and a diverse set of other proteins. Many Hsp90 clients are components of essential signal transduction pathways and regulatory networks, and become only responsive to upstream signals when in complex with Hsp90 or properly folded by Hsp90. Not surprisingly, Hsp90 is involved in many human pathologies, including cancer, neurodegeneration, autoimmune diseases and infections with a wide range of pathogens, and is considered a prime drug target.The aim of this project is to further our understanding of the intricate Hsp90 machinery by (1) performing structure-function analysis of Hsp90-cochaperone complexes and Hsp90-cochaperone-client complexes, (2) screening for novel cochaperones and modifiers of the Hsp90 system necessary for the maturation of kinases in yeast, (3) analyzing the influence of posttranslational modification on conformational dynamics and function of human Hsp90. For our investigations we will use a wide range of biochemical and biophysical methods, including fluorescence techniques, hydrogen exchange mass spectrometry (HX-MS), and protein-protein interaction techniques, and genetics and molecular biology methods in baker’s yeast and mammalian cell culture.