Conformational sub-states in enzyme catalysis: Applications to ribonuclease

酶催化中的构象亚状态：在核糖核酸酶中的应用

• 批准号: 9040996
• 负责人: Pratul K Agarwal
• 金额: $ 50.99万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9040996
• 关键词: Active Sites; Address; Architecture; Biophysics; Catalysis; Communities; Computer Simulation; Data; Data Set; Disease; Environment; Enzyme Kinetics; Enzymes; Eukaryota; Exhibits; Family; Family member; Foundations; Health; Human; Human Genome; Hydrolysis; Individual; Investigation; Joints; Kidney; Knowledge; Life; Medicine; Methodology; Methods; Minor; Modeling; Molecular; Molecular Conformation; Motion; Multienzyme Complexes; Nuclear Magnetic Resonance; Pancreas; Pathogenicity; Physiological; Play; Population; Proteins; RNA; Relaxation; Research; Research Personnel; Resolution; Ribonucleases; Role; Sampling; Shapes; Signal Transduction; Software Framework; Software Tools; Staging; Structure; System; Techniques; Theoretical model; Time; Titrations; Triad Acrylic Resin; United States National Institutes of Health; Validation; Virus; antiangiogenesis therapy; base; chemical reaction; conformer; design; enzyme mechanism; enzyme structure; improved; inhibitor/antagonist; insight; member; millisecond; molecular dynamics; molecular recognition; mutant; neurotoxicity; novel; protein function; protein structure; quantum; research study; scaffold; simulation; software development; theories; tool

DESCRIPTION (provided by applicant): Enzymes perform the designated function of catalyzing chemical reactions by serving more than a scaffold for bringing together the reactants. The role of structure in enzyme function has been known for more than a century now; however, more recent evidence suggests that a functioning enzyme exists in an ensemble of conformations under ambient physiological conditions. The ensemble view of enzyme structure suggests that it can sample conformational sub-states that exhibit function promoting structural and dynamical features. Further, evidence from experiments and computational modeling suggest that transitions between these conformational sub-states enable substrate recognition and catalysis. Quantitative insights into these functionally relevant sub-states remains challenging, particularly due to the wide range of time-scales involved, limited window of resolution for individual techniques and the fact that some of the sub-states can be potentially short-lived. We address these issues by developing a joint computational-experimental framework to identify and characterize such functionally relevant sub-states in the context of enzyme function. In addition to identifying structural intermediates, our framework will quantify the relative population of the conformations in various sub-states as well as enable their linkage to kinetics of enzyme function through the catalytic cycle. This integrated approach will be used to investigate the bio-medically relevant ribonuclease (RNase) family of proteins and enzymes. In particular, we will: (1) Develop a theoretical framework to identify and characterize the multi-scale hierarchy in the conformational landscape of proteins; (2) Utilize the developed framework to investigate the RNase fold members and their ability to access distinct conformational sub-states, including functionally relevant sub-states along the catalytic cycle; (3) Validate the developed model and predicted sub- states by integrating nuclear magnetic resonance (NMR) relaxation dispersion experiments. The developed methodology and models will be improved by iterative interaction between the 3 PIs with different expertise spanning theoretical biophysics, computational simulations and experimental techniques. Overall, our studies will have implications in the design of novel inhibitors of RNase function in the context of neurotoxicity, angiogenesis and anti-pathogenicity.

描述（由申请人提供）：酶通过为将反应物汇总的脚手架提供多的脚手架来执行催化化学反应的指定功能。结构在酶功能中的作用已有一个多世纪以来。但是，最近的证据表明，在环境生理条件下的构象中存在一种功能正常的酶。酶结构的整体视图表明，它可以采样表现出促进结构和动力学特征的功能的构象子态。此外，来自实验和计算建模的证据表明，这些构象子态之间的过渡可以实现底物识别和催化。对这些功能相关的子群的定量见解仍然具有挑战性，特别是由于涉及的时间表很广，单个技术的分辨率有限，并且某些子州可能是可能是可能是潜在的事实 短暂的。我们通过开发一个联合计算实验框架来解决这些问题，以在酶功能的背景下识别和表征此类功能相关的子群。除了识别结构中间体外，我们的框架还将量化各种子群中的构象的相对群体，并通过催化循环使它们与酶功能动力学的联系。这种综合方法将用于研究蛋白质和酶的生物医学相关的核糖核酸酶（RNase）家族。特别是：（1）开发一个理论框架，以识别和表征蛋白质构象局势中的多尺度层次结构； （2）利用开发的框架来研究RNase折叠构件及其访问不同构象子态的能力，包括沿催化循环的功能相关的子群； （3）通过整合核磁共振（NMR）弛豫分散实验来验证开发的模型并预测子态。通过跨越理论生物物理学，计算模拟和实验技术的不同专业知识之间的3个PI之间的迭代相互作用将改善开发的方法和模型。总体而言，我们的研究将在神经毒性，血管生成和抗病原体的背景下对新型RNase功能抑制剂的设计具有影响。