The 3.0 angstrom resolution structure of Flock House virus and its functional implications
Abstract
Icosahedral viral capsids built from a single gene product are among the simplest biological assembly systems, yet the details of their assembly, stability, and disassembly are poorly understood. The Nodaviruses are among the simplest animal viruses known. They are small, spherical, non-enveloped viruses that infect insects, mice, and fish. Their capsid is constructed of 180 copies of a single gene product that assembles into a T = 3 quasi-equivalent lattice. When T is larger than one, strict symmetry cannot be maintained in subunit packing. Therefore, a subunit must be capable of assembling into pentamers and hexamers. Previous high resolution structures of viral capsids displaying quasi-equivalence show a portion of the capsid subunit mediating the different interactions by the order or disorder of a peptide arm. The structure of Flock House nodavirus (FHV), discussed in this thesis, revealed that the nucleic acid genome, along with a peptide arm from the capsid protein, modulates the different subunit contacts to assemble a T = 3 quasi-equivalent lattice. FHV propagated from infected Drosophila cells, crystallizes into single crystals that diffract X-rays. The 3.0A resolution crystal structure of Flock House virus was determined. The structure revealed 20 nucleotides ordered at each icosahedral 2-fold axis forming a helical duplex. The RNA interacts with a helical protein domain of the subunit that lies inside the contiguous coat shell. One of the helices that binds the RNA is part of a 44 amino acid polypeptide that is autocatalytically cleaved from the initial subunit translation product after virion assembly. The structure suggests that RNA associated with the cleaved polypeptide may be important in the infection process. The maturation cleavage is necessary for virus infectivity. Therefore, a competent foreign expression system is required to investigate mutants of the capsid protein. A procedure was developed to express FHV capsid protein in a baculovirus system. A residue at the cleavage site was mutated to prevent virus maturation, and the expressed protein spontaneously assembles into virus-like particles. Crystals were grown of these mutant provirions that diffract X-rays beyond 3.3A resolution.
Degree
Ph.D.
Advisors
Johnson, Purdue University.
Subject Area
Biophysics|Microbiology
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