Small spherical viruses are among the simplest replicating systems in biology, yet factors affecting their architecture, assembly, stability, disassembly and interactions with their hosts are still poorly understood. These problems will be investigated with icosahedral viruses displaying two types of quasisymmetry; the T=3 nodaviridae (180 identical subunits) and T=4 tetraviridae (240 identical subunits). They will be investigated by X-ray crystallography, various biophysical methods, molecular genetics and methods of computational chemistry. The concepts derived from these basic studies may guide the investigation of complex medically important viruses displaying quasi equivalence. Primary and tertiary structural studies of nodaviruses and the tetravirus, nudaurelia omega capensis virus (N-omega-V), suggest that they probably share assembly and maturation strategies as well as a mechanism for regulating quasi-symmetry, although the final capsid types are substantially different. The detailed structure function-relationship governing architecture, assembly, stability, and infectivity will be analyzed for each system separately and for both systems comparatively. A wealth of information has been developed over the last decade for both virus groups and this allows the specific aims described below a strong chance of success. Nodaviridae (T=3): (1) The structure of Flock House virus (FHV) will be refined. (2) An infectuous clone of FHV is available. In addition the expression of the coat protein gene of FHV in the baculovirus system results in the spontaneous assembly of virus like particles and these have been purified and crystallized in the laboratory of the P.I. A variety of site-directed mutations designed to test hypotheses of infectivity, assembly and stability will be generated with one or both of these systems and they will be characterized by biological, biophysical, and computational chemistry methods and, when, appropriate, by crystallography. 3) The structure of Nodamuravirus will be determined and refined and factors affecting its different stability and host range, compared to other nodaviruses, will be explored. Tetraviridae (T=4): (1) The high resolution structure of N-omega-V will be completed and refined. (2) Expression of the cloned coat protein gene of N-omega-V in baculovirus will be attempted and the system will be used for mutagenesis studies to test hypotheses based on the structure. Expressed mutants of N-omega-V will be characterized as described for the nodaviridae (3) The gene sequence of NOV polymerase and the entire gene of another tetravirus, N-beta-V, will be determined.
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