Mutations in the hydrophobic core and in the protein-RNA interface affect the packing and stability of icosahedral viruses.

The information required for successful assembly of an icosahedral virus is encoded in the native conformation of the capsid protein and in its interaction with the nucleic acid. Here we investigated how the packing and stability of virus capsids are sensitive to single amino acid substitutions in the coat protein. Tryptophan fluorescence, bis-8-anilinonaphthalene-1-sulfonate fluorescence, CD and light scattering were employed to measure urea- and pressure-induced effects on MS2 bacteriophage and temperature sensitive mutants. M88V and T45S particles were less stable than the wild-type forms and completely dissociated at 3.0 kbar of pressure. M88V and T45S mutants also had lower stability in the presence of urea. We propose that the lower stability of M88V particles is related to an increase in the cavity of the hydrophobic core. Bis-8-anilinonaphthalene-1-sulfonate fluorescence increased for the pressure-dissociated mutants but not for the urea-denatured samples, indicating that the final products were different. To verify reassembly of the particles, gel filtration chromatography and infectivity assays were performed. The phage titer was reduced dramatically when particles were treated with a high concentration of urea. In contrast, the phage titer recovered after high-pressure treatment. Thus, after pressure-induced dissociation of the virus, information for correct reassembly was preserved. In contrast to M88V and T45S, the D11N mutant virus particle was more stable than the wild-type virus, in spite of it also possessing a temperature sensitive growth phenotype. Overall, our data show how point substitutions in the capsid protein, which affect either the packing or the interaction at the protein-RNA interface, result in changes in virus stability.