Accounting for Conformational Dynamics in Post-Translational Phosphorylation of Signaling Proteins

解释信号蛋白翻译后磷酸化的构象动力学

• 批准号: 1616741
• 负责人: Jeffrey Peng
• 金额: $ 51.32万
• 依托单位: University of Notre Dame
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-15 至 2021-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1616741&HistoricalAwards=false
• 关键词: Accounting; Conformational; Dynamics; Post; Translational

Title: Accounting for Conformational Dynamics in Post-Translational Phosphorylation of Signaling ProteinsTo thrive, cells must regulate dense networks of protein-protein interactions that drive basic processes such as cell growth, division, and death. One well-known regulatory mechanism is "post-translational phosphorylation" - the reversible addition of phosphate groups to specific amino acids within a protein. Phosphorylation can alter protein binding specificity, enzymatic activity, or sub-cellular location, thereby regulating its interaction with other proteins. But just how this comes about is not yet clear. In fact, an atomic level explanation for how site-specific phosphorylation changes a protein's interaction properties remains an open challenge. This project addresses this challenge by investigating the effects of post-translational phosphorylation on protein conformational dynamics. The protein segments subject to phosphorylation are often the same segments involved in intermolecular interactions. Critically, they often have conformational flexibility that defies explanation by standard biochemical analysis. By investigating how these dynamic protein segments convert phosphorylation into functional change, this project will provide insight into a widespread, yet poorly understood aspect of protein phosphorylation. Gaining this insight is a necessary step toward understanding and manipulating protein interaction networks. This project will bring interdisciplinary educational opportunities to students at early stages of college by developing freshmen courses that introduce physical principles based on biophysical phenomena. This project will also train doctoral students and undergraduate research interns, including members of underrepresented groups, to become highly qualified scientists in the area biophysics and computational biology; the cultivation of such expertise is vital to keep the U.S. competitiveness in research and in the global economy.The project will combine liquid-state multi-dimensional Nuclear Magnetic Resonance (NMR) spectroscopy, molecular dynamics (MD) simulations, and mutagenesis to map how site-specific phosphorylation impacts protein dynamics and protein-substrate interactions in the mitotic signaling protein, Pin1. Pin1 interactions with other proteins are sensitive to both conformational dynamics and post-translational phosphorylation. This proposal will determine the extent to which phosphorylation exploits electrostatic networks to achieve long-range site-to-site communication (allostery). This project will also define how phosphorylation of intrinsically disordered regions nevertheless elicit functional changes at remote binding sites. Because Pin1 interacts with numerous other signaling proteins, results of this project will give broad insight into the mechanisms by which dynamic protein segments convert phosphorylation into functional change. This project will also include development of general protein NMR methods to better characterize dynamic electrostatic interactions in proteins, and their description in terms of conformational ensembles. This project is supported by Molecular and Cellular Biosciences Division in the Directorate for Biological Sciences.

标题：解释信号蛋白翻译后磷酸化的构象动力学细胞为了茁壮成长，必须调节密集的蛋白质-蛋白质相互作用网络，这些网络驱动着细胞生长、分裂和死亡等基本过程。一种众所周知的调节机制是“翻译后磷酸化”--磷酸基团可逆地加到蛋白质中的特定氨基酸上。磷酸化可以改变蛋白质的结合特异性、酶活性或亚细胞位置，从而调节其与其他蛋白质的相互作用。但这究竟是如何发生的还不清楚。事实上，对特定位点的磷酸化如何改变蛋白质的相互作用性质的原子水平解释仍然是一个悬而未决的挑战。这个项目通过研究翻译后磷酸化对蛋白质构象动力学的影响来解决这一挑战。被磷酸化的蛋白质片段往往是参与分子间相互作用的相同片段。关键的是，它们往往具有无法用标准生化分析解释的构象灵活性。通过研究这些动态蛋白质片段如何将磷酸化转化为功能变化，该项目将提供对蛋白质磷酸化的一个广泛但鲜为人知的方面的洞察。获得这一洞察力是理解和操纵蛋白质相互作用网络的必要步骤。该项目将通过开发介绍基于生物物理现象的物理原理的新生课程，为大学早期阶段的学生带来跨学科的教育机会。该项目还将培训博士生和本科生实习生，包括代表不足的群体的成员，成为生物物理学和计算生物学领域的高素质科学家；培养此类专业知识对于保持美国在研究和全球经济中的竞争力至关重要。该项目将结合液态多维核磁共振(核磁共振)谱、分子动力学(MD)模拟和突变，绘制特定位点的磷酸化如何影响有丝分裂信号蛋白Pin1中的蛋白质动力学和蛋白质-底物相互作用。Pin1与其他蛋白质的相互作用对构象动力学和翻译后磷酸化都很敏感。这一提议将决定磷酸化在多大程度上利用静电网络实现远距离的点对点通信(变构)。该项目还将定义内在无序区域的磷酸化如何引起远程结合位点的功能变化。由于Pin1与许多其他信号蛋白相互作用，该项目的结果将使人们对动态蛋白片段将磷酸化转化为功能变化的机制有更广泛的了解。该项目还将包括开发通用的蛋白质核磁共振方法，以更好地表征蛋白质中的动态静电相互作用，并根据构象系综来描述它们。该项目得到了生物科学局分子和细胞生物科学司的支持。