Towards the structure determination of Coxsackie virus B3
Abstract
Coxsackie virus B3 (CVB3) is a small, icosahedral, single-stranded RNA virus belonging to the enterovirus genus within the picornavirus family. The enteroviruses include poliovirus, the coxsackie A and B viruses, hepatitis A virus and the echoviruses. The coxsackie B viruses are causative agents of a wide variety of mild to severe diseases and are the most common agents known to cause viral myocarditis. CVB3 can cause myocarditis in mice and provides an excellent animal model in which to study virus structure-function and drug therapy. CVB3 crystallizes in a primitive monoclinic space group (a = 574.6A, b = 302.1A, c = 521.6A, $\beta$ = 107.7$\sp\circ$) with two virions in the asymmetric unit. The crystals diffract well to 2.7A resolution and 66% of the X-ray diffraction data have been collected to 3.0A resolution. Systematically weak reflections and the self-rotation function confirms pseudo R32 symmetry. This pseudo R32 symmetry orients and positions each particle in the monoclinic cell near face-centered positions. The resultant packing arrangement is equally consistent with space groups P2 or P2$\sb1$. Establishing the deviations from R32 packing for both independent particles in the monoclinic asymmetric unit and determining the true space group (P2 or P2$\sb1$) has been the most challenging aspect of this project. These parameters must be accurately known in order to refine and extend phases to the resolution limits of the available data and to successfully utilize the high order of non-crystallographic redundancy (120-fold) during molecular replacement real space averaging. A series of crystal soaking experiments were performed with human rhinovirus 14 (HRV14) using subfragments of antiviral compounds (Win compounds). It was observed that these small Win compounds, at high concentrations, bind into the $\beta$-barrel pocket of VP1 and cause conformational changes similar to those of the larger Win compounds. It was also observed that dimethyl sulfoxide (DMSO), the solvent for the Win compounds, binds specifically to HRV14 at high concentrations. One of the DMSO molecules binds into the pocket near the pore opening and the other two bind outside the pocket in the canyon near the protomer-protomer interface.
Degree
Ph.D.
Advisors
Rossmann, Purdue University.
Subject Area
Biochemistry|Microbiology
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