Manipulating and predicting the unfolded ensembles of disordered proteins

操纵和预测无序蛋白质的未折叠整体

• 批准号: 10224244
• 负责人: Patricia Louise Clark
• 金额: $ 34.95万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10224244
• 关键词: Address; Adopted; Affect; Amino Acid Sequence; Amino Acids; Behavior; Cell physiology; Charge; Collaborations; Complex; Computer Models; Computing Methodologies; Consensus; Data; Data Analyses; Deuterium; Development; Dimensions; Disease; Evolution; Fluorescence Resonance Energy Transfer; Foundations; Geometry; Goals; Gram-Negative Bacteria; Hour; Huntington Disease; Hydrogen; Hydrogen Bonding; Hydrophobicity; In Vitro; Knowledge; Lead; Measurement; Measures; Mediator of activation protein; Membrane; Methods; Modeling; Molecular; Molecular Conformation; Mutation; Pattern; Physiological; Play; Positioning Attribute; Procedures; Property; Protein Secretion; Proteins; Radial; Research; Roentgen Rays; Role; Science; Solvents; Stretching; Structure; Sum; Surface; System; Testing; Vertebral column; Virulence; Walking; Water; amyloid formation; biophysical techniques; experimental study; improved; in vivo; innovation; maltose-binding protein; molecular recognition; non-Native; novel; periplasm; pertactin; physical property; polyglutamine; polypeptide; predictive modeling; preference; protein folding; protein function; simulation; tool

PROJECT SUMMARY The physical properties of intrinsically disordered proteins (IDPs) affect their positive (functional) and negative (disease-causing) roles in cell function. Yet despite intense effort, we still lack a predictive understanding of the physical properties that govern whether a given polypeptide chain sequence will adopt an expanded or collapsed conformational ensemble under physiological conditions – and by extension, how collapse affects protein function. The Sosnick and Clark labs have formed a collaboration to develop precisely this understanding. This project consists of experimental studies of IDPs tightly integrated with new data analysis procedures and computational modeling tools. This project builds on our recent findings with “PNt”, a 334 residue IDP that under physiological conditions adopts an expanded ensemble of conformations well approximated by a self-avoiding random walk, despite having an amino acid content that, according to the current paradigm, should lead to a collapsed, self-associated state. We hypothesize that current models fail to predict the behavior of PNt due to current knowledge gaps regarding which sequence patterns, beyond global sequence composition, lead to collapse. We propose that deviations from an expanded state, e.g., adopting a collapsed globule, are due to specific sequence patterns including local stretches of hydrophobic residues. We will test this hypothesis by reordering (“shuffling”) the amino acid sequence of PNt to induce collapse, and likewise shuffle the amino acid sequence of maltose binding protein (MBP) to promote expansion of its denatured state, which we recently demonstrated is highly collapsed. We will use a battery of biophysical methods (SAXS, hydrogen-deuterium exchange, NMR) to measure the extent of local versus global collapse, assessing the sensitivity of conformational ensembles to subtle changes in sequence order, and the relationship between collapse and hydrogen bonding. We will determine which hydrophobicity scales yield the best predictions of experimental results for polypeptide chain collapse, and use the effects of shuffling to parameterize simulations. Finally, we will test the impact of altering IDP collapse on protein function in vivo, specifically the efficient secretion of autotransporter virulence proteins to the surface of Gram-negative bacteria. Our overall goal is to accurately predict the conformational ensemble for a user-inputted amino acid sequence and solvent condition.

项目概要 本质无序蛋白质 (IDP) 的物理特性影响其积极（功能）和消极 然而，尽管付出了巨大的努力，我们仍然缺乏对细胞功能的预测性理解。 控制给定多肽链序列是否采用扩展或扩展的物理特性 生理条件下塌陷的构象集合 - 以及塌陷如何影响 索斯尼克和克拉克实验室已经合作开发了这一功能。 该项目包括与新数据分析紧密结合的国内流离失所者实验研究。 第 334 章 残基IDP在生理条件下能够很好地采用扩展的构象集合 通过自回避随机游走来近似，尽管氨基酸含量根据 当前的范式应该导致崩溃的、自我关联的状态，我们追求的是当前的模型无法实现。 由于当前关于哪些序列模式的知识差距，超出全局范围，预测 PNt 的行为 我们建议偏离扩展状态，例如采用 塌陷的小球是由于特定的序列模式（包括疏水性残基的局部延伸）造成的。 将通过重新排序（“改组”）PNt 的氨基酸序列以诱导崩溃来检验这一假设，并且 同样地改组麦芽糖结合蛋白（MBP）的氨基酸序列以促进其扩展 我们最近证明了变性状态是高度塌陷的，我们将使用生物物理电池。 方法（SAXS、氢-氘交换、NMR）来测量局部与整体塌陷的程度， 评估构象系综对序列顺序细微变化的敏感性，以及 我们将确定哪种疏水性尺度会产生崩溃和氢键之间的关系。 多肽链塌陷实验结果的最佳预测，并利用改组的效果 最后，我们将测试改变 IDP 崩溃对体内蛋白质功能的影响， 特别是自转运蛋白毒力蛋白有效分泌到革兰氏阴性菌表面 我们的总体目标是准确预测用户输入的氨基酸的构象集合。 顺序和溶剂条件。

项目成果

Water as a Good Solvent for Unfolded Proteins: Folding and Collapse are Fundamentally Different.

水作为未折叠蛋白质的良好溶剂：折叠和塌陷是根本不同的。

• 发表时间: 2020
• 影响因子: 5.6
• 作者: Clark, Patricia L;Plaxco, Kevin W;Sosnick, Tobin R
• 通讯作者: Sosnick, Tobin R

How hydrophobicity, side chains, and salt affect the dimensions of disordered proteins.

疏水性、侧链和盐如何影响无序蛋白质的尺寸。

• 发表时间: 2024-05
• 作者: Baxa, Michael C;Lin, Xiaoxuan;Mukinay, Cedrick D;Chakravarthy, Srinivas;Sachleben, Joseph R;Antilla, Sarah;Hartrampf, Nina;Riback, Joshua A;Gagnon, Isabelle A;Pentelute, Bradley L;Clark, Patricia L;Sosnick, Tobin R
• 通讯作者: Sosnick, Tobin R

Development of in vivo HDX-MS with applications to a TonB-dependent transporter and other proteins.

开发体内 HDX-MS 并应用于 TonB 依赖性转运蛋白和其他蛋白质。

• 发表时间: 2022-09
• 作者: Lin, Xiaoxuan;Zmyslowski, Adam M;Gagnon, Isabelle A;Nakamoto, Robert K;Sosnick, Tobin R
• 通讯作者: Sosnick, Tobin R

Prediction and Validation of a Protein's Free Energy Surface Using Hydrogen Exchange and (Importantly) Its Denaturant Dependence.

使用氢交换及其（重要的）其变性依赖性来预测和验证蛋白质的自由能表面。

• 发表时间: 2022-01-11
• 作者: Peng, Xiangda;Baxa, Michael;Faruk, Nabil;Sachleben, Joseph R;Pintscher, Sebastian;Gagnon, Isabelle A;Houliston, Scott;Arrowsmith, Cheryl H;Freed, Karl F;Rocklin, Gabriel J;Sosnick, Tobin R
• 通讯作者: Sosnick, Tobin R