Capturing Transient Protein and Nucleic Acid Structures During Their Functions on Multiple Spatial and Temporal Scales

捕获在多个空间和时间尺度上发挥作用期间的瞬时蛋白质和核酸结构

• 批准号: 10264031
• 负责人: Lin X Chen
• 金额: $ 35.04万
• 依托单位: NORTHWESTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10264031
• 关键词: Active Sites; Antineoplastic Agents; Binding; Biochemical Reaction; Biological; Biological Process; Calcium ion; Catalysis; Computing Methodologies; Consumption; Coupled; Crystallization; DNA Structure; Data; Data Analyses; Data Collection; Deposition; Diffusion; Drug Delivery Systems; Drug Design; Electromagnetic Fields; Environment; Enzymes; Evolution; Future; Health; Human; Investigation; Kinetics; Laboratories; Laboratory Research; Lasers; Ligands; Light; Lipids; Maps; Measures; Membrane; Metabolism; Metals; Methodology; Methods; Microfluidic Microchips; Modeling; Modification; Molecular; Molecular Conformation; Molecular Structure; Motion; Nucleic Acid Folding; Nucleic Acids; Oncogenes; Optics; Oxidation-Reduction; Phase; Phase Transition; Physiologic pulse; Physiological Processes; Protein Conformation; Protein Dynamics; Proteins; Pump; RNA Conformation; Reaction; Regulation; Research; Resolution; Respiration; Roentgen Rays; Sampling; Scientist; Signal Transduction; Source; Spectrum Analysis; Stimulus; Structure; Synchrotrons; System; Temperature; Time; Translations; Work; X ray diffraction analysis; X ray spectroscopy; X-Ray Crystallography; absorption; biological systems; biomacromolecule; chromophore; cytochrome c oxidase; design; emission spectroscopy; environmental change; improved; inhibitor/antagonist; innovation; insight; instrumentation; macromolecule; milligram; millisecond; molecular dynamics; nanocarrier; nanometer; novel; novel strategies; nucleic acid structure; programs; protein data bank; protein folding; protein structure; response; simulation; three dimensional structure; time use; tool; x-ray free-electron laser

Summary/Abstract The long term objective of the proposed research is to develop an integrated instrumentation capable of studying protein/nucleic acid structural dynamics that are relevant to their functions on the time scales from femtosecond to millisecond in order to gain new insight into correlations of active site structures and global conformations of these molecules. Snapshots of solution phase molecular structures over different spatial scales, from sub-Ångström for active sites to several nanometers for overall conformation, will be captured using time-resolved X-ray spectroscopy and scattering. These structural studies will be combined with advanced molecular dynamics simulations that will generate detailed atomistic dynamics consistent with measured scattering profiles over a wide-range of temporal scales from femtosecond to millisecond. The proposed research is complementary to single crystal X-ray diffraction, and intends to map reaction trajectories through three-dimensional structures as a function time in media that mimic biological environments. In order to detect structural changes in an ensemble, reaction triggers must be designed to create sudden environmental changes that synchronize actions of the molecules with much higher time resolution than traditional mixing. The program has three main innovations from previous studies: 1) to develop triggering sources beyond direct light excitation used in the past to initiate reactions to overcome the limitation that very few biological systems related to human health are light activated for their function; 2) to develop novel sample delivery system that reduces the sample consumption by a factor of 100 and enables many precious laboratory samples to be studied using the time-resolved X-ray methods; and 3) to develop a combined approach in data analyses using advanced molecular dynamics simulation coupled to time-dependent X-ray scattering data to extract structures with improved structural accuracy especially for those coexisting species. The above innovation in methodology will allow us to investigate a number of systems that are biologically significant for enzymatic reactions, signal sensing, protein/nucleic acid folding/unfolding as well as lipids phase transitions. Several systems are chosen for the proposed studies to capture transient structures of, a) local metal center and global protein conformations of cytochrome c oxidase model proteins triggered by photodissociation of inhibitors; b) protein folding induced by calcium ion a concentration jump; c) temperature-induced RNA conformational changes sensing signal for translation; d) pH-dependent DNA structures for human oncogene regulation and e) pH-responsive lipid nanocarrier assembly for anticancer drug delivery. These structural results combined with those of reaction kinetics from optical transient spectroscopy will provide guidance for modulating protein and nucleic acid functions via structural modifications, which will lead to impacts in drug design, enzymatic function enhancement, catalysis, as well as theoretical calculations.

摘要/摘要 拟议研究的长期目标是开发一种能够 在时间尺度上研究与其功能相关的蛋白质/核酸结构动力学 从飞秒到毫秒，以获得对活动中心结构和 这些分子的全球构象。不同温度下溶液相分子结构的快照 空间尺度，从活性中心的亚纳斯特伦到总构象的几个纳米，将是 使用时间分辨X射线光谱分析和散射进行捕获。这些结构研究将结合在一起 先进的分子动力学模拟将产生与详细的原子动力学一致的 在从飞秒到毫秒的大范围时间尺度上测量的散射分布。 建议的研究是对单晶X射线衍射的补充，并打算绘制反应图 在模拟生物的介质中作为时间函数的三维结构的轨迹 环境。为了检测合奏中的结构变化，反应触发器必须设计成 创造突然的环境变化，使分子的行动与更长的时间同步 分辨率高于传统混合。该计划在以往研究的基础上有三个主要创新：1)到 开发超越过去使用的直接光激发的触发来源，以启动反应，以克服 与人类健康有关的极少数生物系统因其功能而被光激活的局限性； 2)开发新颖的样本运送系统，将样本消耗量减少100倍，以及 使许多珍贵的实验室样本能够用时间分辨的X射线方法进行研究；以及3) 使用高级分子动力学模拟开发数据分析的组合方法 用含时X射线散射数据提取结构，特别是提高了结构精度 对于那些共存的物种。上述方法上的创新将使我们能够调查一些 对酶反应、信号传感、蛋白质/核酸具有重要生物学意义的系统 折叠/展开以及脂类相变。为拟议的研究选择了几个系统。 为了捕捉，a)细胞色素c的局部金属中心和整体蛋白构象的瞬时结构 抑制剂光解离引发的氧化酶模型蛋白：b)钙诱导的蛋白质折叠 离子a浓度跃迁；c)温度诱导的RNA构象变化传感信号 D)人类癌基因调节的pH依赖的DNA结构和e)pH响应性脂质 用于抗癌药物输送的纳米载体组件。这些结构结果与反应的结果相结合 来自光学瞬变光谱的动力学将为蛋白质和核酸的调控提供指导 通过结构修饰发挥作用，这将对药物设计、酶功能产生影响 增强，催化，以及理论计算。