CHEETAH Center for the Structural Biology of HIV Infection, Restriction, and Viral Dynamics

核心摘要 该核心旨在为切达中心成员提供新的成像和测量方法 我们中心正在研究的不同HIV宿主系统的行为。具体而言，我们的结构和 机制研究核心（核心2），将开发新的方法，并提供切塔中心成员获得 在荧光光谱学和动力学、结构支架方面拥有最先进的仪器和专业知识 测定和电子冷冻断层扫描。 组成部分1（荧光光谱学和动力学）将提供荧光方面的仪器和专门知识 标记和单分子荧光方法来研究生物结构和动力学。的新方法 将生物过程与真实的时间相关联，并将cryoEM结构研究与动态分析联系起来， 开发 组分2（用于结构确定的支架）将产生并提供称为“合成的”的抗体样分子。 抗原结合物”（sAB）作为cryoEM结构测定的基准标记。该部分还将优化和 应用新的、授权的“通用”分子基准类别，其基于：1）可转运的结合表位，或2） 第二抗体对于更具挑战性的RNA和RNA-蛋白质复合物，也正在创建通用基准。 组件3（电子冷冻断层扫描）将结合相关的强大和快速发展的技术， 光学和电子显微镜（CLEM），聚焦离子束（FIB）铣削，和电子冷冻断层扫描（ECT）重建 以前所未有的分辨率呈现自然分子景观。该组件还将开发新的功能相关 可以通过荧光或通过电子能量损失光谱（EELS）成像定位的报告分子，并应用这些 报告人在HIV-1亚病毒组织，病毒进入，病毒核心复制和贩运的问题。

CORE SUMMARY This Core is designed to provide CHEETAH Center members with access to novel approaches for imaging and measuring the behaviors of different HIV-host systems under investigation in our Center. Specifically, our Tools for Structural and Mechanistic Studies Core (Core 2), will develop new methodology and provide CHEETAH Center members with access to state-of-the-art instrumentation and expertise in Fluorescence Spectroscopy and Dynamics, Scaffolds for Structure Determination, and Electron Cryotomography. Component 1 (Fluorescence Spectroscopy and Dynamics) will provide instrumentation and expertise in fluorescence labeling and single-molecule fluorescence approaches to studies of biological structure and dynamics. Novel methods for correlating biological processes in real time and for linking cryoEM structural studies to dynamic analyses are also being developed. Component 2 (Scaffolds for Structure Determination) will generate and provide antibody-like molecules called “Synthetic Antigen Binders” (sABs) as fiducial markers for cryoEM structure determinations. The Component will also optimize and apply new, empowering classes of “universal” molecular fiducials based on: 1) transportable binding epitopes, or 2) secondary antibodies. Universal fiducials for more challenging RNAs and RNA-protein complexes are also being created. Component 3 (Electron Cryotomography) will incorporate the powerful and rapidly advancing technologies of correlative light and electron microscopy (CLEM), focused ion beam (FIB) milling, and electron cryotomography (ECT) to reconstruct native molecular landscapes at unprecedented resolution. The Component will also develop new functional correlative reporters that can be located by fluorescence or by electron energy-loss spectroscopy (EELS) imaging, and apply these reporters to problems in HIV-1 sub-viral organization, virus entry, and viral core replication and trafficking.