Decoding the molecular logic of TPR cochaperones

项目总结/摘要 三肽重复序列（TPR）辅伴侣蛋白是一个不同的衔接蛋白家族，与转录因子相互作用， 热休克蛋白（Hsp）70和Hsp 90分子伴侣系统。这些模块化蛋白质由它们的 表位TPR结构域，其介导与Hsps的复合物形成，以及酶或支架 领域，可以帮助监护人在客户招聘或质量控制。这些配件的重要性 在维持蛋白质稳态的功能是证明了TPR共伴侣蛋白的影响，在广泛的 从神经退化到癌症的一系列疾病。然而，我们对分子机制的理解 这些蛋白质识别底物的基础是不完整的。特别是，越来越多的证据表明， TPR结构域可以募集超出其典型伴侣的蛋白质， 这种替代途径目前尚不清楚。此外，通常难以建立底物 考虑到蛋白质稳态内可能发生的补偿， 网络遗传变异。可特异性抑制TPR的化学探针的研制 因此，具有高时间分辨率的辅助分子伴侣复合物是非常需要的。努力发展这样的 通过了解TPR结构域的哪些分子特征可以 用于实现高亲和力和选择性结合。该提案的目的是开发一种化学品， 一个工具包，通过解码TPR之间相互作用的分子逻辑，帮助解决这两个问题 辅伴侣及其底物。在我的第一个目标中，我将开发化学蛋白质组学工具来分析 的TPR cochaperones和测量的底物协会在不同的刺激响应的变化。这些 探针还将使得TPR抑制剂的特异性能够以快速的方式进行评估。在我的第二个目标，我 将改进预测评分功能，以便全面识别自主 由E3连接酶CHIP识别。在探索CHIP与其相互作用的分子决定因素时， 底物，我也将确定的功能，使一个单独的TPR辅伴侣结合配体是不同的 从其相关的家庭成员。这项工作是重要的，因为它将提供基本的见解， 规则的TPR共分子伴侣的生物学，并将作为未来的底物发现的基石 以及在这个蛋白质家族中的探针开发工作。拟议的研究还将提供培训， 环境非常适合我成为一名翻译化学生物学家的目标，有机会 在培养研究能力的同时，我也获得了最先进技术的经验 独立地。

PROJECT SUMMARY / ABSTRACT Tetratricopeptide repeat (TPR) cochaperones are a diverse family of adaptor proteins that cooperate with the heat shock protein (Hsp) 70 and Hsp90 chaperone systems. These modular proteins are composed of their eponymous TPR domain, which mediates complex formation with the Hsps, and an enzymatic or scaffolding domain that can aid the chaperone in client recruitment or quality control. The importance of these accessory functionalities in maintaining protein homeostasis is evidenced by the implication of TPR cochaperones in a wide range of diseases from neurodegeneration to cancer. However, our understanding of the molecular mechanisms that underpin substrate recognition by these proteins is incomplete. In particular, there is increasing evidence that TPR domains can recruit proteins beyond their canonical chaperone binding partners, and the significance of this alternative pathway is currently unknown. In addition, it is generally difficult to establish substrate relationships for TPR cochaperones given the compensation that can occur within the protein homeostasis network following genetic perturbations. Development of chemical probes that can specifically inhibit TPR cochaperone complexes with high temporal resolution is therefore highly desirable. The effort to develop such tools would be greatly augmented by an understanding of which molecular features of TPR domains can be exploited to achieve high affinity and selective binding. The objective of this proposal is to develop a chemical toolkit that helps solve both of these problems by decoding the molecular logic of interactions between TPR cochaperones and their substrates. In my first aim, I will develop chemical proteomic tools to profile the binding of TPR cochaperones and measure changes in substrate association in response to different stimuli. These probes will also enable the specificity of TPR inhibitors to be assessed in a rapid fashion. In my second aim, I will refine a predictive scoring function in order to comprehensively identify substrates that are autonomously recognized by the E3 ligase CHIP. In exploring the molecular determinants of CHIP's interactions with its substrates, I will also identify features that enable an individual TPR cochaperone to bind ligands that are distinct from its related family members. This work is significant because it will provide fundamental insights into the rules governing the biology of TPR cochaperones, and will serve as the bedrock for future substrate discovery and probe development efforts across this protein family. The proposed studies will also provide a training environment that is ideally suited to my goal of becoming a translational chemical biologist, with opportunities to gain experience with state-of-the-art techniques while also cultivating my ability to conduct research independently.