Crystallographic and biophysical analyses of isometric viruses will be performed to study virus stability, protein-nucleic acid interactions, the interactions of antibodies with viruses, and the evolution of spherical viruses. The subjects of our study will be comoviruses and nepoviruses. Both these virus groups share many properties with the animal picornaviruses. Results from the previous funding period have shown that at least one of the nucleoprotein components of the comovirus, beanpod mottle virus (BPMV), has 20% of the RNA genome organized with icosahedral symmetry. A total of 11 contiguous ribonucleotides were modeled in the icosahedral asymmetric unit of 3482 nucleotides. This structure will be refined at 2.8 angstrom resolution and structures of the empty capsid and the bottom component (containing 5889 nucleotides) will be determined and refined. A third component, which is biologically identical to the bottom component but has exchanged endogenous polyamines for cesium ions, will also be investigated. Crystals of all the components are isomorphous and different to 2.6 angstrom resolution. Collaboration arrangements with an immunologist have been made to obtain monoclonal antibodies to cowpea mosaic virus (CPMV) and BPMV. Attempts will be made to co-crystallize Fab fragments with the virus particles. An extraordinary crystal form of CPMV, in which FAB fragments have been successfully diffused, will also be studied. Collaborations with a molecular biologist have been made to study, by site directed mutagenesis, factors affecting assembly, protein- nucleic acid interactions and the effect of specific changes in the coat protein on the host specificity in comoviruses. The structure of tobacco ringspot virus (TRV) will be studied using x-ray crystallography. This nepovirus is similar to comoviruses, but it contains one capsid proteins of 58kD. Given its similarity to comoviruses in size, component types, and genome sizes, it may have ordered RNA and subunit is probably composed of 3 covalently linked beta-barrel domains.
