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

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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.