STRUCTURE OF VIRAL RNA AND ITS ROLE IN ASSEMBLY

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DESCRIPTION (provided by applicant): A variety of physical and chemical techniques, chief among them being X-ray crystallography, will be applied to a series of icosahedral viruses with the ultimate objectives of (1) deducing the detailed structure of the RNA genomes inside the viruses, (2) identifying the important protein-nucleic acid interactions, and (3) devising models for the assembly and disassembly of the virions consistent with existing molecular biological data. Viruses to be studied are Satellite Tobacco Mosaic Virus (STMV), Satellite Tobacco Necrosis Virus (STNV), Satellite Panicum Mosaic Virus (SPMV), Turnip Yellow Mosaic Virus (TYMV), Desmodium Yellow Mottle Virus (DYMV), Tomato Aspermy Virus (TAV), Brome Mosaic Virus (BMV), Panicum Mosaic Virus (PMV), and empty capsids of each of the last five species. These are viruses whose structures are best known, for which RNA is at least partially visible in the virion, and for which the greatest amount of experience exists. Core RNA particles will be prepared by digestion of the intact viruses and will be crystallized for X-ray diffraction analyses. Empty capsids will be made, crystallized, and their diffraction data used in difference Fourier syntheses to reveal RNA structure. With integration of the proposed chemical and enzymatic probe information, and predictions from computer modeling to supplement the X-ray studies, a comprehensive reconciliation of structure with function will emerge. We have shown that when atomic force microscopy is applied in conjunction with chemical and enzymatic reagents, complex viruses can be dissected to reveal internal components, including nucleic acid. To extend our investigations of nucleic acid conformation and packaging to larger, more complex viruses we will extend our AFM investigations of the internal structures of HIV, herpes simplex virus, mouse leukemic virus, and vaccinia virus. This research is significant because the conformation of the nucleic acid component of no virus is currently known, nor is there any comprehensive structural basis for the processes of viral assembly and disassembly.