Cryo EM/ET Core

冷冻 EM/ET 核心

• 批准号: 10506949
• 负责人: Peijun Zhang
• 金额: $ 33.88万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10506949
• 关键词: Area; Attention; Capsid; Cells; Classification; Complement; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Electron Microscopy; Environment; Fluorescence Microscopy; HIV; HIV-1; Image; In Situ; Individual; Infection; Integration Host Factors; Ions; Label; Macromolecular Complexes; Methods; Microscopy; Nuclear Import; Nuclear Pore; Nuclear Pore Complex; Play; Process; Proteins; Resolution; Sampling; Scanning Electron Microscopy; Software Tools; Structure; Techniques; Technology; Viral; Virion; Work; X-Ray Crystallography; density; experience; flexibility; glycoprotein 41; inhibitor; light microscopy; nanometer resolution; new technology; nuclear pore complex protein 96; particle; protein complex; single molecule; small molecule inhibitor; software development; trafficking

Cryo-EM/ET Core Cryo-EM and cryo-ET are powerful techniques for high-resolution structural studies of proteins and protein complexes, complementing X-ray crystallography and NMR methods. Cryo-ET with subtomogram averaging and classification, in particular, has recently gained abundant attention as a prime method for characterization of macromolecular complexes that are intrinsically flexible and often challenging to purify for structure determination by single particle cryo-EM. Moreover, in situ cryo-ET can inform on structures of complexes within their native cellular environment, with near atomic resolution. The Cryo-EM/ET Core will play a key role in developing and applying these cutting-edge methods for structural analysis of heterogeneous HIV-1 viral components and their complexes with host cell factors and small molecule inhibitors. The Core will support PCHPI projects by providing structural information on assembly and maturation intermediates as well as capsid-host interactions during nuclear import and integration. Furthermore, the Core will offer new capabilities of cryo-EM and cryo-ET imaging of infectious samples in a biosafety level 3 (BSL-3) environment and develop new technologies and workflows for single-molecule correlation between cryo-ET and super-resolution fluorescence microscopy. These approaches will yield transformative structural and mechanistic information into the key steps of HIV-1 replication.

CRYO-EM/ET核心 冷冻EM和冷冻-ET是用于蛋白质和蛋白质高分辨率结构研究的强大技术 复合物，补充X射线晶体学和NMR方法。具有亚图平均和 特别是分类最近引起了广泛的关注，作为表征的主要方法 大分子络合物本质上是柔韧性的，通常具有挑战性，以确定结构 由单个粒子冷冻EM。此外，原位冷冻-ET可以告知其当地中的复合物结构 细胞环境，近乎原子分辨率。 Cryo-Em/et核心将在发展和 应用这些尖端方法，用于异质HIV-1病毒成分的结构分析及其 具有宿主细胞因子和小分子抑制剂的复合物。核心将通过提供PCHPI项目 有关组装和成熟中间体以及在山皮相互作用的结构信息 核进口和整合。此外，核心将提供Cryo-Em和Cryo-ET成像的新功能 生物安全3级（BSL-3）环境中的传染性样品并开发新技术和工作流程 用于冷冻-ET和超分辨率荧光显微镜之间的单分子相关性。这些 方法将产生变革性的结构和机械信息，成为HIV-1的关键步骤 复制。