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.

低温EM/ET堆芯 CRYO-EM和CRYO-ET是研究蛋白质和蛋白质高分辨率结构的有力技术 配合物，补充X射线结晶学和核磁共振方法。亚断层图像平均和亚断层图像平均的冷冻机 尤其是分类，作为一种主要的表征方法，最近得到了广泛的关注。 大分子络合物本质上是灵活的，而且经常具有结构确定的提纯难度 通过单粒子冷冻-EM。此外，原位冷冻-ET还可以了解其天然配合物的结构 蜂窝环境，具有接近原子分辨率。Cryo-EM/ET核心将在开发和 应用这些尖端方法分析HIV-1病毒异质组分及其结构 与宿主细胞因子和小分子抑制剂的复合体。该核心将通过以下方式支持PCHPI项目 关于组装和成熟中间体以及衣壳与宿主相互作用的结构信息 核进口和核电一体化。此外，核心将提供低温电磁和冷冻-ET成像的新能力 生物安全3级(BSL-3)环境中的感染样本，并开发新技术和工作流程 用于低温电子显微镜和超分辨荧光显微镜之间的单分子关联。这些 方法将在艾滋病毒-1的关键步骤中产生变革性的结构性和机械性信息 复制。