Structure Determination Technology (592-634)

结构测定技术 (592-634)

• 批准号: 9352608
• 负责人: WAYNE A. HENDRICKSON
• 金额: $ 4.93万
• 依托单位: NEW YORK STRUCTURAL BIOLOGY CENTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9352608
• 关键词: Address; Algorithmic Analysis; Algorithms; Characteristics; Classification; Cluster Analysis; Collection; Complex; Cryoelectron Microscopy; Crystallization; Crystallography; Data; Data Analyses; Data Collection; Data Quality; Data Set; Dehydration; Development; Electrons; Elements; Environment; Generations; Harvest; Helium; Ion Channel; Ligand Binding; Ligands; Maps; Measurement; Membrane; Membrane Proteins; Methionine; Methods; Modeling; Molecular Conformation; Noise; Pathology; Phase; Photons; Play; Population; Production; Radiation induced damage; Resolution; Roentgen Rays; Role; Selenomethionine; Signal Transduction; Structure; Synchrotrons; System; Technology; Time; Ursidae Family; absorption; base; design; detector; electron density; experience; experimental study; improved; innovation; new technology; novel; particle; prevent; protein structure; synchrotron radiation; x-ray free-electron laser

Atomic structures of membrane proteins are playing a major role in understanding the mechanisms of action and deriving fundamental principles for these systems. Crystallography is the dominant method for obtaining membrane protein structures at atomic and near atomic resolutions, single-wavelength anomalous diffraction (SAD) predominates for de novo structure determination, which is essential in many instancs. Recently x-ray free electron lasers for crystallography and direct electron detectors for single-particle cryo electron microscopy (cryo-EM) emerging as technologies for solving challenging structures of membrane proteins and their complexes. Nevertheless, bottlenecks remain including weak anomalous signals, micron-sized crystals and radiation damage in crystallography and atomic interpretation of electron density maps from cryo-EM. We propose to address these bottleneck challenges in four sub-projects. Aim 1 (TR&D 4.1) focuses on the extraction of signals from weak anomalous scatterers with the aim of making native-SAD analysis robust even for low-resolution membrane protein structures. Aim 2 (TR&D 4.2) relates to challenges in the manipulation and analysis of microcrystals, including the development of new methods for mounting microcrystals presentation in microdiffraction experiments and methods of analysis for optimal synthesis of complete data sets from the partial data of many crystals. Aim 3 (TR&D 4.3) is concerned with the mitigation of radiation damage, which is exacerbated in measurements from microcrystals as needed for optimal native-SAD analyses. Aim 4 (TR&D 4.4) will address the optimized fitting of atomic models to cryo-EM maps. We will innovate methods for identifying specific residues types in such maps and methods for a sorted classification of ligand-binding populations for optimized placement of the ligands and associated conformational changes.

膜蛋白的原子结构在理解作用机制方面起着重要作用， 这些系统的基本原理。结晶学是获得膜的主要方法 原子和近原子分辨率的蛋白质结构，单波长异常衍射（SAD） 主要用于从头结构测定，这在许多情况下是必要的。最近x射线自由电子 用于晶体学的激光器和用于单粒子低温电子显微镜（cryo-EM）的直接电子探测器 作为解决膜蛋白及其复合物的挑战性结构的技术。然而，尽管如此， 瓶颈仍然存在，包括微弱的异常信号，微米尺寸的晶体和晶体学中的辐射损伤 和低温电镜电子密度图的原子解释。我们建议解决这些瓶颈挑战 四个子项目。目标1（TR&D 4.1）侧重于从弱异常散射体中提取信号， 目的是使天然SAD分析即使对于低分辨率膜蛋白结构也是稳健的。目标2（TR&D 4.2） 涉及操作和分析微晶的挑战，包括开发新的方法， 微衍射实验中的安装微晶演示和优化合成的分析方法 从许多晶体的部分数据得到完整的数据集。目标3（TR&D 4.3）涉及减少 辐射损伤，这是加剧了从微晶测量所需的最佳native-SAD 分析。目标4（TR&D 4.4）将解决原子模型与冷冻EM图的优化拟合问题。我们将创新 用于鉴定这种图谱中的特定残基类型的方法和用于配体结合的分类的方法 用于优化配体的放置和相关的构象变化的群体。