A positively charged surface patch on the pestivirus NS3 protease module plays an important role in modulating NS3 helicase activity and virus production

Pestivirus nonstructural protein 3 (NS3) is a multifunctional with protease and helicase activities that are essential for virus replication. In this study, we used combination of biochemical genetic approaches to investigate the relationship between positively charged patch on module NS3 function. The surface composed four basic residues, R50, K74 K94 in domain H24 structurally integrated cofactor NS4APCS. Single-residue or simultaneous four-residue substitutions alanine aspartic acid had little effect ATPase activity. However, single substitution resulted apparent changes activity RNA-binding ability NS3. When these mutations were introduced into classical swine fever (CSFV) cDNA clone, at (Qua_A Qua_D) impaired production infectious virus. Furthermore, replication efficiency CSFV variants was partially correlated vitro. Our results suggest conserved plays an important role modulating vitro production.