Conformational dynamics, binding and aggregation of intrinsically disordered proteins

总结 内含子无序蛋白（IDP）占真核生物蛋白质组的30%以上。他们执行至关重要的 细胞中的信号传导、转录和翻译等功能，它们调节和控制细胞 周期它们的功能失常导致了一些最具挑战性的疾病，这对美国的健康越来越重要。 护理系统，如癌症和神经变性。这些疾病的相关费用是一些 在美国增长最快，增长最快。国内流离失所者在病毒的复制和传播中也起着关键作用， 病原体为了正常发挥作用，IDP必须（i）有效地结合特异性结合伴侣;和（ii）避免 病理性聚集在分子水平上理解IDP序列如何编码这两个 过程，以及细胞中的突变和压力条件如何影响它们，将大大促进我们的研究。 理解和治疗和预防这些疾病的能力。 目前主要的试验性努力集中在：（一）解决国内流离失所者的结构问题， 结合机制（IDP结构/功能）;或（ii）将IDP聚集机制解析为IDP 聚集成特定的淀粉样蛋白聚集体（IDP功能障碍）。在这里，我们提出了一个完全不同的，物理的 化学方法研究IDP构象动力学对结合机制的影响; 以及IDP相分离与聚集之间的定量关系。我们将使用高 分辨率的实验技术和方法，在我们的实验室，沿着多尺度模拟。 由于大多数IDP通过偶联折叠和结合结合，因此预期它们的构象动力学将 大大影响了绑定机制。我们将IDP构象动力学纳入结合的方法 研究将提供一个关键的缺失环节，以了解IDP的功能结合。 我们提出的研究建立在一组IDP的结合机制的最新特征， 重点研究1）构象动力学对结合的影响; 2） 液相分离及其与病理性聚集的联系。我们将联合收割机结合不同的高分辨率 实验技术-包括纳秒激光泵浦光谱-与分子模拟， 描述国内流离失所者的结构和动态。新的方法将被用来量化液相分离， 国内流离失所者。目标I的结果：通过比较不同结合的IDP动力学来检验我们的假设 目标II：通过扰动序列和溶剂调节结合机制， III：量化疾病突变对构象动力学、相分离和微结构的影响。 FUS_LC的聚集;将对卵巢癌中涉及的IDP的分子理解产生直接影响。 和乳腺癌，在副氧化病毒的复制和肌萎缩侧索硬化症， 额颞叶痴呆这些结果有可能改变我们看待耦合折叠的方式 和结合以及液相分离，这些对于IDP功能至关重要。

Summary Intrinsically disordered proteins (IDPs) make up more than 30% of eukaryotic proteomes. They carry out vital functions in the cell, such as signaling, transcription and translation, and they regulate and control the cell cycle. Their malfunction leads to some of the most challenging diseases, of growing concern to the US health care system, such as cancer and neurodegeneration. The associated cost of these diseases are some of the highest and fastest growing in the US. IDPs also play a key role in the replication and spreading of viral pathogens. In order to function properly IDPs must (i) bind efficiently to specific binding partners; and (ii) avoid pathological aggregation. A molecular level understanding of how IDP sequences encode for these two processes, and how mutations and stress conditions in cells can affect them, will significantly advance our understanding and ability to treat and prevent such diseases. Major experimental efforts are currently focused on: (i) resolving the structural ensembles of IDPs and the binding mechanism (IDP structure/function); or (ii) resolving the mechanisms of IDP aggregation into IDP aggregation into specific amyloid aggregates (IDP malfunction). Here we propose a radically different, physical chemistry approach in which we study the effect of IDP conformational dynamics on the binding mechanism; and the quantitative relation between IDP phase separation and aggregation. We will do so by using high resolution experimental techniques and methods developed in our lab, along with the multiscale simulations. Because most IDPs bind through coupled folding and binding, their conformational dynamics is expected to greatly affect the binding mechanism. Our approach of incorporating IDP conformational dynamics in binding studies will provide a key missing link to understand IDP functional binding. Our proposed study builds on the recent characterization of the binding mechanism of a group of IDPs, and focuses on studying 1) the effect of conformational dynamics on binding; and 2) the physiological process of liquid phase-separation and its link to pathological aggregation. We will combine different high resolution experimental techniques -including nanosecond laser pump spectroscopy- with molecular simulations to characterize IDP structure and dynamics. Novel methods will be used to quantify liquid phase-separation of IDPs. Results from Aim I: Test our hypothesis by comparing IDP dynamics for different binding scenarios; Aim II: Modulating the binding mechanism by perturbing the sequence and solvent, and Aim III: Quantify the effect of a disease mutation on the conformational dynamics, phase separation and aggregation of FUS_LC; will have direct impact on the molecular understanding IDPs implicated in ovarian and breast cancer, in the replication of paramoxyviruses and in amyotrophic lateral sclerosis and frontotemporal dementia. These results have the potential of transforming our way of viewing coupled folding and binding, and liquid phase separation which are crucial for IDP function.