X-Ray Determination of Proteins and Viruses

蛋白质和病毒的 X 射线测定

• 批准号: 9603571
• 负责人: Michael Rossmann
• 金额: $ 41.82万
• 依托单位: Purdue Research Foundation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-02-15 至 2000-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9603571&HistoricalAwards=false
• 关键词: Ray; Determination; Proteins; Viruses

9603571 Rossmann X-ray diffraction, using crystallized viruses, will be the primary tool to analyze the assembly and uncoating intermediates of various viruses. Frequently, however, it is not possible to crystallize transient intermediates on account of their short-lived, unstable properties. In these cases, the X-ray crystallography will be augmented by cryo electron microscopy. The major emphasis will be on the study of bacterial viruses, such as single-stranded X174 and the far more complex double-stranded T4. The far more complex and far larger dsDNA T4 phage has over 40 structural proteins. While it is unlikely that it will ever be possible to study the whole virus by X-ray diffraction, it is possible to study the atomic structure of individual proteins. Combination of X-ray diffraction and cryo-EM is to be used to determine the interactions of components. Part of the work will be to develop suitable techniques for interfacing cryoelectron microscopy with X-ray diffraction techniques and also to examine particles that lack perfect icosahedral symmetry. The objectives will be to determine not only the structures, but also the molecular mechanisms by which the assembly intermediates are stabilized and triggered into the subsequent steps of morphogenesis. Similarly, the structural intermediates of viral disassembly during cellular infection are to be examined. Comparision of different viral systems should lead to the identification of common strategies and evolutionary relationships. The goal of the research is to elucidate the mechanisms by which infectious virus particles are assembled from their component parts. Although the work is devoted to the study of bacterial viruses, the viral mechanisms are also likely to be utilized by plant and animal viruses. The different process of disassembly during viral entry into a host will also be studied. Viral assembly often requires scaffolding proteins which help guide protein folding. Hence, the stud ies will be among the first to observe the structures of these chaperone-like proteins complexed with their substrates. The study of their transient intermediates will require the development of techniques to combine X-ray crystallography (which can resolve features near atomic resolution) and cryo-electron microscopy (which has poorer resolution but lacks the requirement of making crystals) Combination of these techniques is likely to be critical in the future development of structural biology.

9603571 Rossmann X 射线衍射使用结晶病毒，将成为分析各种病毒的组装和脱壳中间体的主要工具。 然而，由于瞬态中间体的寿命短、性质不稳定，通常不可能使其结晶。 在这些情况下，X 射线晶体学将通过冷冻电子显微镜得到增强。 主要重点是细菌病毒的研究，例如单链 X174 和更复杂的双链 T4。 更为复杂和更大的 dsDNA T4 噬菌体具有 40 多种结构蛋白。 虽然不可能通过 X 射线衍射研究整个病毒，但研究单个蛋白质的原子结构是可能的。 X 射线衍射和冷冻电镜的结合将用于确定组分的相互作用。 部分工作将是开发合适的技术，将冷冻电子显微镜与 X 射线衍射技术结合起来，并检查缺乏完美二十面体对称性的粒子。 目标不仅是确定结构，还确定组装中间体稳定并触发后续形态发生步骤的分子机制。 同样，还需要检查细胞感染期间病毒分解的结构中间体。 不同病毒系统的比较应该导致共同策略和进化关系的识别。 该研究的目的是阐明传染性病毒颗粒从其组成部分组装的机制。 尽管这项工作致力于研究细菌病毒，但病毒机制也可能被植物和动物病毒利用。 还将研究病毒进入宿主期间的不同分解过程。 病毒组装通常需要支架蛋白来帮助指导蛋白质折叠。 因此，这些研究将是首批观察这些类伴侣蛋白与其底物复合的结构的研究之一。 对它们的瞬态中间体的研究将需要开发结合X射线晶体学（可以解析接近原子分辨率的特征）和冷冻电子显微镜（分辨率较差但缺乏制造晶体的要求）的技术。这些技术的结合可能对结构生物学的未来发展至关重要。