Poliovirus mutants at histidine 195 of VP2 do not cleave VP0 into VP2 and VP4

The final stage of poliovirus assembly is characterized by a cleavage of th e capsid precursor protein VP0 into VP2 and VP4. This cleavage is thought t o be autocatalytic and dependent on RNA encapsidation. Analysis of the poli ovirus empty capsid structure has led to a mechanistic model for WO cleavag e involving a conserved histidine residue that is present in the surroundin g environment of the VP0 cleavage site. Histidine 195 of VP2 (2195H) is hyp othesized to activate local water molecules, thus initiating a nucleophilic attack at the scissile bond. To test this hypothesis, 2195H mutants were c onstructed and their phenotypes were characterized. Consistent with the req uirement of VP0 cleavage for poliovirus infectivity, all 2195H mutants were nonviable upon introduction of the mutant genomes into HeLa cells, Replace ment of 2195H with threonine or arginine resulted in the assembly of a high ly unstable 150S virus particle. Further analyses showed that these particl es contain genomic RNA and uncleaved VP0, criteria associated with the prov irion assembly intermediate. These data support the involvement of 2195H in mediating VP0 cleavage during the final stages of virus assembly.