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

核心摘要 此核心旨在为猎豹中心成员提供使用成像和测量的新方法 在本中心接受调查的不同HIV宿主系统的行为。具体地说，我们用于结构和 机械研究核心(核心2)，将开发新的方法，并为猎豹中心成员提供访问 在荧光光谱和动力学、结构支架方面的最先进仪器和专业知识 测定和电子冷冻断层摄影术。 组成部分1(荧光光谱和动力学)将提供荧光方面的仪器和专业知识 生物结构和动力学研究的标记和单分子荧光方法。新的方法可用于 将生物过程实时关联起来，并将低温EM结构研究与动态分析联系起来，也正在进行中 发展起来的。 组件2(用于结构确定的支架)将产生并提供抗体样分子，称为“合成的 抗原结合剂“(SABS)作为低温电子显微镜结构测定的基准标记物。该组件还将优化和 基于：1)可运输的结合表位，或2)，应用新的、赋予能力的“通用”分子基准类别。 二次抗体。更具挑战性的RNA和RNA-蛋白质复合体的通用基准也正在创建中。 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)球磨和电子冷冻断层扫描(ECT)重建 以史无前例的分辨率绘制原生分子景观。该组件还将开发新的功能关联 可以通过荧光或电子能量损失谱(EELS)成像定位的记者，并应用这些 记者在HIV-1亚病毒组织、病毒进入以及病毒核心复制和贩运方面存在问题。

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.