Structural protein complexes sedimenting at 140S, 70S (empty capsids), and 14S were isolated from foot-and-mouth disease virus-infected cells. The empty capsids were stable, while 14S complexes were relatively short-lived. Radioimmune binding assays involving the use of neutralizing monoclonal antibodies to six distinct epitopes on type A12 virus and polyclonal antisera to A12 structural proteins demonstrated that native empty capsids were indistinguishable from virus. Infected cell 14S particles possessed all the neutralizing epitopes and reacted with VP2 antiserum. Cell-free structural protein complexes sedimenting at 110S, 60S, and 14S containing capsid proteins VP0, VP3, and VP, are assembled in a rabbit reticulocyte lysate programmed with foot-and-mouth viral RNA. These structures also contain the six epitopes, and cell-free 14S structures like their in vivo counterparts reacted with VP2 antiserum. Capsid structures from infected cells and the cell-free complexes adsorbed to susceptible cells, and this binding was inhibited, to various degrees, by saturating levels of unlabeled virus. These assays and other biochemical evidence indicate that capsid assembly in the cell-free system resembles viral morphogenesis in infected cells. In addition, epitopes on the virus surface possibly involved in interaction with cellular receptor sites are found early in virion morphogenesis.
Initial experiments in our laboratory have successfully reassembled infectious polyoma virions from dissociated virion products. Virions treated with ethyleneglycol-bis-N,N'-tetraacetic acid and the reducing agent ,B-mercaptoethanol at pH 7.5 were dissociated to a 48S DNA-protein complex and capsomere subunits. The virion dissociation products were not infectious by plaque assay and lacked hemagglutination activity. These virion dissociation products were reassembled to intact virions by overnight dialysis against a reassembly buffer containing CaCl2, dimethyl sulfoxide, and Triton X-100 in phosphate-buffered saline at pH 7.4. The biophysical characteristics of the reassembled virions were identical to those of untreated virions in that the reassembled virions had a sedimentation value of 240S in sucrose gradients and a buoyant density of 1.315 g/cm: in CsCl isopycnic gradients. The reassembled virions were intact as determined by electron microscopy and were found to be 60% resistant to DNase I treatment. Biologically, the reassembled purified virions were found to partially regain both hemagglutinating activity and plaque-forming ability.
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