Technology Development 1: Development of nanocrystal processing protocols for microED

技术开发1：microED纳米晶体加工方案的开发

• 批准号: 9977961
• 负责人: Guillermo Alberto Calero
• 金额: $ 29.74万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-27 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9977961
• 关键词: Binding; Biocompatible Materials; Biological; Catalogs; Communities; Crystallization; Crystallography; Development; Drops; HIV; HIV-1; Human; Laboratories; Methodology; Methods; Micro Electron Diffraction; Molecular Biology; Multiprotein Complexes; Positioning Attribute; Program Development; Property; Protein Chemistry; Proteins; Protocols documentation; Resolution; Sampling; Selection Criteria; Speed; Structural Biologist; Structural Protein; Structure; Synchrotrons; System; Technology; Time; Transmission Electron Microscopy; Universities; X-Ray Crystallography; beamline; detector; electron crystallography; free-electron laser; infancy; macromolecule; meter; nanocrystal; nanometer; novel; prevent; protein complex; protein structure; technology development; x-ray free-electron laser

Abstract for Technology Development Program 1 - MicroED X-ray crystallography over the last 50 years has been the most successful method to obtain atomic structures of biological molecules. Crystallization of large multi-protein complexes is challenging, and even if successful, the resulting crystals are usually small, delicate and present multiple challenges. New developments such as serial femtosecond crystallography using a free electron laser (FEL) and electron crystallography or micro-electron diffraction (microED) are alternative approaches to obtain structures, using nano-meter or low micro-meter sized crystals (nanocrystals). Solving structures of bio-macromolecules using FELs requires billions of nanocrystals, amounts of material that frequently is not available. Solving structures with microED, on the other hand, may, in principle, be possible with only a few nanocrystals, thus overcoming problems related to sample quantity. The Calero laboratory at the University of Pittsburgh has pioneered nanocrystal discovery and optimization using transmission electron microscopy (TEM). This has permitted to significantly increase the number of potential crystallization conditions through direct observation of crystal lattices. In this technology development proposal, we intend to develop new methodologies to determine structures of protein complexes of HIV-1 proteins and their human binding partners by exploring nano crystallization space and optimizing stabilizing conditions for the application of microED approaches for structure determination.

技术开发计划摘要1-微型 X射线晶体学在过去50年中一直是获得原子结构的最成功的方法 生物分子。大型多蛋白络合物的结晶是具有挑战性的，即使成功， 产生的晶体通常很小，细腻，并且出现了多个挑战。诸如系列的新发展 飞秒晶体学使用游离电子激光（FEL）和电子晶体学或微电子 衍射（微型）是使用纳米米或低微米获得结构的替代方法 大小的晶体（纳米晶体）。使用FELS解决生物大分子的结构需要数十亿美元 纳米晶体，经常不可用的材料量。用微型解决结构，在 另一方面，原则上可能只有几个纳米晶体可能是可能的，从而克服了与 样品数量。匹兹堡大学的Calero实验室已经开创了纳米晶体发现和 使用透射电子显微镜（TEM）优化。这已允许显着增加 通过直接观察晶格的潜在结晶条件的数量。在这项技术中 开发建议，我们打算开发新方法来确定蛋白质复合物的结构 通过探索纳米结晶空间并优化HIV-1蛋白及其人类结合伙伴 用于应用微型方法进行结构确定的稳定条件。