This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. We are analyzing immunotherapeutics and vaccines for use against several viral hemorrhagic fevers including Ebola virus, Dengue virus, and Lassa Fever virus. A major complication to development of these medical countermeasures is that in many instances, we do not understand where on the viral surface glycoproteins the most effective antibodies bind. For some viruses, it appears that the angle at which the antibody binds the virus will determine efficacy, as many epitopes are sterically hidden from immune recognition. Although high-resolution crystal structures are tremendously informative in defining sites and angles of interaction, not all of these antibodies will crystallize in complex with their cognate glycoprotein, and we have fifty+ antibodies of interest to map. In addition, many of the antibodies against the Ebola virus are directed against a heavily glycosylated mucin-like domain which we can model by SAXS but cannot crystallize. We would like to use SAXS to roughly map binding sites of our panels of antibodies, using known crystal structures of antibodies and unbound glycoproteins as molecular models to fit SAXS scattering. SAXS models will be confirmed with biochemical data. Our preliminary data using this technique to map antibody sites against both Dengue and Ebola point to a fruitful line of inquiry ahead For example, we have been able to localize binding of key antibodies to certain faces and domains of the antigens and support these models with mutagenesis and peptide binding studies.
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