Proteolytic activity of the cowpea mosaic virus encoded 24K protein synthesized in Escherichia coli.

The function of the 24-kilodalton (24K) protein encoded by cowpea mosaic virus (CPMV) has been studied by constructing a bacterial expression plasmid that contained a cloned chimeric segment consisting of partial DNA copies of CPMV M-RNA (including sequences coding for both capsid proteins) and B-RNA (including sequences coding for the 24K protein). Viral sequences were transcribed from the phage T7 promoter phi 10 of plasmid pT7-6 using T7-RNA polymerase expressed from plasmid pGP1-2 present in the same cells. Upon inducing the synthesis of T7-RNA polymerase several new polypeptides that contained CPMV-specific sequences were expressed, as demonstrated by immunoprecipitation and immunoblotting. Furthermore a proteolytic activity was detected in induced cells which cleaved the viral protein sequences specifically at two glutamine-glycine sites. One of the cleavage products represented capsid protein VP23. The proteolytic activity was absent when an 87-bp deletion was introduced in the coding region for the 24K protein, indicating that this protein represented the protease involved in the proteolytic processing at those specific sites.