Impact of charge regulation on conformational and phase equilibria of intrinsically disordered proteins

细胞被组织成不同的隔间，其中许多隔间由不同的酸度定义。如果细胞中的蛋白质含有可电离的氨基酸，那么富含质子的环境可以改变蛋白质上的电荷。由质子的结合或解离引起的蛋白质电荷状态的变化可以控制细胞中的蛋白质如何对特定类型的信号做出反应，例如引起盐或质子浓度变化的环境因素。该项目的研究将利用新的方法来研究特定的蛋白质，即所谓的固有无序蛋白质，是如何通过吸收或释放基于溶液酸度的质子来修饰的。作为该项目的一部分进行的研究将有助于更好地理解蛋白质电荷状态的变化是如何影响其功能的。这项工作很重要，因为电荷介导的相互作用是各种蛋白质功能的核心，包括一种蛋白质对另一种蛋白质的识别。鉴于推动该项目研究的技术进步的独特性质，在分子生物物理学的广泛领域中，计算和实验的交叉点将有新的培训机会。该项目的发现将被纳入本科生的课程材料。此外，参与该项目的关键人员将与市中心初中和高中的学生密切合作，解释pH值和对酸碱平衡的更好理解将如何推动该项目出现的新研究前沿。计算和实验的结合，在一定程度上是由包括机器学习模块在内的新型计算工具的发展推动的，将为解决分子生物物理学中具有挑战性的问题创造新的工作流程。这些模块将被纳入分子生物物理学研究实习生和学生的教学模块。该项目的目标是了解可电离残基之间的质子交换和这些残基周围溶液离子的积累如何有助于内在无序蛋白质(IDPs)的电荷状态和构象。目前，我们对IDPs的形式、功能和相行为的理解受到简化的假设的限制，即可电离残基的电荷状态由模型化合物的pKa值固定。拟议的工作基于构象波动会导致可电离残基的局部微环境发生变化的原理。这些局部异质性可以诱导可滴定位置之间的质子交换。这也会导致溶液离子在可电离残基周围优先积累。推动这些研究的一个关键进展是Q正则系综的最新发展，它是一种基于正式和严格的统计物理描述电荷状态和构象异质性的联合效应的方法。重要的是，已经开发了一个集成的实验和计算管道，这使得人们能够利用Q正则系综结构来量化质子缔合或解离的影响，即所谓的电荷调节，以及它通过电荷重整化与溶液离子的相互作用。基于电荷测量和构象测量之间形式解耦的方法，将允许将来自电位滴定和构象或相平衡测量的信息纳入到模拟中。当以实验数据为基础时，这将导致对在不同溶液条件下可访问的电荷状态和构象的系综的完整描述。这些方法是新颖的，重点是独特的，从该项目中产生的见解可能是前所未有的。该项目由分子和细胞生物科学部的分子生物物理组资助。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Cells are organized into distinct compartments, and many of these compartments are defined by different acidity. An environment rich in protons can change the charge on proteins in cells if the proteins have amino acids that are ionizable. Changes to charge states of proteins that arise from the association or dissociation of protons can control how proteins in cells respond to specific types of signals such as environmental factors that induce changes to salt or proton concentrations. Research in this project will leverage new methods to study how specific proteins, known as intrinsically disordered proteins, are modified by the uptake or release of protons based on the acidity of the solution. Studies carried out as part of the project will lead to an improved understanding of how alterations to charge states of proteins influence their functions. This work is important because charge-mediated interactions are central to a variety of protein functions including the recognition of one protein by another. Given the unique nature of the technological advances that are driving research in the project, there will be novel training opportunities at the intersection of computation and experiment in the broad area of molecular biophysics. Findings from the project will be incorporated into materials of courses for undergraduates. Further, key personnel who are involved in the project will work closely with students from inner city middle and high-schools to explain how pH and a better understanding of acid-base equilibria will drive new research frontiers that emerge from the project. The blend of computation and experiment, driven in part by the development of novel computational tools, including machine learning modules, will create new workflows for solving challenging problems in molecular biophysics. These modules will be incorporated into instructional modules for research trainees and students in molecular biophysics.The goal of the project is to understand how the exchange of protons between ionizable residues and the accumulation of solution ions around these residues contribute to the charge states and conformations of intrinsically disordered proteins (IDPs). Currently, our understanding of the form, functions, and phase behaviors of IDPs is limited by the simplifying assumption that charge states of ionizable residues are fixed by the pKa values of model compounds. The proposed work rests on the principle that conformational fluctuations engender changes to local microenvironments of ionizable residues. These local heterogeneities can induce exchange of protons among titratable sites. This can also induce preferential accumulation of solution ions around ionizable residues. A key advance that drives the investigations is the recent development of the q-canonical ensemble, which is a formal and rigorous statistical physics-based description of the joint effects of charge state and conformational heterogeneity. Importantly, an integrated experimental and computational pipeline has been developed and this allows one to leverage the structure of the q-canonical ensemble to quantify the effects of proton association or dissociation, known as charge regulation, and its interplay with the effects of solution ions through charge renormalization. Approaches that rest on the formal decoupling between measurements of charge and conformation, will allow for the incorporation of information derived from potentiometric titrations and measurements of conformational or phase equilibria into simulations. This, when anchored by experimental data, will lead to a full description of the ensemble of charge state and conformations that are accessible at different solution conditions. The approaches are novel, the focus is unique, and the insights to be generated from the project are likely to be unprecedented. This project is funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.