Poxviruses are a family of large DNA viruses that encode up to 200 distinct open reading frames.The large size of the poxvirus genome is an important feature that has allowed them to acquiremultiple immunomodulatory genes and thereby evolve unique strategies for evasion from host antiviralresponses. Ectromelia virus (EV) is a member of the orthopoxvirus family and is a highlyvirulent rodent pathogen that causes the disease mousepox. EV is similar to variola virus, thecausative agent of human smallpox. Our primary hypothesis is that secreted and cell membraneassociated proteins encoded by EV likely serve important roles in viral evasion of host mediatedinnate and adaptive immune responses. Using a bioinformatics approach coupled to theestablished literature, we have selected 28 target proteins from the EV Moscow strain genome thatwill be investigated by a combination of biochemical, functional, and crystallographic tools in a highthroughput,structural genomics style approach. Our primary targets of investigation include theseven known cytokine and chemokine decoy receptors encoded by the virus that are specific forTNE CD30L, IL-18, IFN-alpha, IFN-gamma, IL-lbeta, and CC-chemokines. We are also targetingthree proteins with sequence similarity to natural killer receptors of the C-type lectin family. Wehave the following specific aims for the exploration of these potential agents of immune subterfuge:(1) Establish baculovirus and bacterial oxidative refolding expression systems for targeted EVencoded proteins to be used in functional and structural studies; (2) Identify and characterize theinteractions between EV proteins and their host ligands and receptors; (3) Determine the structuralbasis of EV protein function by x-ray crystallography and structure-based mutagenesis.
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