MIRA: Uncover Design Rules for Interaction and Assembly of Nature’s Molecular Machines

项目摘要/摘要： 史无前例的高分辨率生物构建块结构的出现正在给 我们可以使蛋白质机制合理化的方法，然而这种结构只提供了一个必要的，但不充分的起点 指向揭示蛋白质的性质、活性和功能。在理解上有一个基本的知识鸿沟 将蛋白质的结构和表面化学转化为蛋白质的性质和功能。即使是像这样的财产 被广泛研究的蛋白质溶解度不能根据已知的蛋白质结构根据我们目前的情况进行先验预测 工具和知识，正如许多单一突变引起最小结构突变的观察所强调的那样 在极大地改变了稳定性、性能和可聚集性的同时，也发生了变化。我们的愿景是揭开这些代码 用于将蛋白质表面结构特性转化为蛋白质表面活性、相互作用和功能。汉人 Lab正在致力于实现这种翻译，其中包括借助先进的光谱方法来探测 局部蛋白质动力学、特定部位的水合特性和构象集合。这些测量结果 由现有的最先进的工具实现，如电子顺磁共振(EPR)线形 分析、脉冲偶极电子顺磁共振和固体核磁共振以及新的方法 由HAN实验室开发，如Overhauser动态核极化(ODNP)等DNP扩增 核磁共振方法。这些方法的结合将使得能够检测蛋白质表面的水合、拓扑 和相互作用，在生理条件下的稀溶液状态下，具有增强的灵敏度。韩氏实验室 将系统地应用和改进研究蛋白质稳定性、相互作用、相分离、 从齐聚到聚合。五年的目标集中在以下精选类别的蛋白质上。我们选择 球状蛋白LOV作为揭示结构-功能关系的模型来辅助合理的蛋白质 导致增强结合亲和力、变构、荧光性质和受控性质的工程 构象可塑性。我们选择tau蛋白来研究一种本质上无序的蛋白(Idp)。 单点突变和微妙的合作伙伴交互显著调整了其稳定性和聚合倾向 在疾病的背景下。我们的目标将是揭示机制，例如通过水化扰动或构象 整体变化，通过这种变化，突变和其他修饰调节聚集倾向。最后，我们 目的揭示寡聚化的结构和机理基础以及功能后果 两种跨膜蛋白，PR和A2a。这项提案的长期目标是破译设计规则 对于蛋白质的相互作用和活性表面，所以从蛋白质表面的结构和拓扑结构可以 预测结合位点的位置，或者学习如何设计合理调节蛋白质-蛋白质组装的结合位点， 并选择或设计特定的聚合路径。

Project Summary/Abstract: The emergence of unprecedented high-resolution structures of biological building blocks is revolutionizing the way we can rationalize the protein machinery, yet the structure only offers a necessary, but insufficient, starting point to uncover protein property, activity and function. There is a fundamental knowledge gap in understanding the translation of protein structure and surface chemistry into protein property and function. Even a property as widely studied as protein solubility cannot be a priori predicted from a known protein structure with our current tools and knowledge, as underscored by the observation that many single mutations invoke minimal structural changes while dramatically changing the stability, property and aggregability. Our vision is to uncover the code for translating protein surface structural properties into protein surface activity, interaction and function. The Han lab is working on achieving such translation, among others, aided by advanced spectroscopy methods that probe local protein dynamics, site-specific hydration properties and conformational ensembles. These measurements are enabled by existing state-of-the-art tools, such as electron paramagnetic resonance (EPR) lineshape analysis, pulsed dipolar EPR and solid-state nuclear magnetic resonance (NMR), as well as novel approaches developed by the Han lab, such as Overhauser dynamic nuclear polarization (ODNP) and other DNP-amplified NMR methods. The combination of these methods will enable the detection of protein surface hydration, topology and interaction, in dilute solution state under physiological conditions and with enhanced sensitivity. The Han lab will systematically apply and refine the tools and methods to study protein stability, interaction, phase separation, oligomerization to aggregation. The five-year goals focus on the following select classes of proteins. We choose the globular protein LOV as a model to uncover the structure-function relationship to aid in rational protein engineering leading to properties such as enhanced binding affinity, allostery, fluorescent property and controlled conformational plasticity. We choose the protein tau to study an intrinsically disordered protein (IDP) for which single-point mutations and subtle partner interactions significantly tune its stability and aggregation propensity in disease contexts. Our goal will be to reveal the mechanisms, e.g. by hydration perturbation or conformational ensemble shifts, through which mutations and other modifications modulate aggregation propensity. Finally, we aim to uncover the structural and mechanistic basis, as well as functional consequences, of oligomerization of two trans-membrane proteins, PR and A2A. The long-term goal of this proposal is to decipher the design rules for interactions and active surfaces of proteins, so that from the protein surface structure and topology one can predict where the binding site is, or learn how to design one that rationally modulates protein-protein assembly, and select or design specific aggregation pathways.