Insights into the oligomerization state–helicase activity relationship of West Nile virus NS3 NTPase/helicase

Abstract West Nile virus (WNV) is a member of the Flaviviridae family of positive-strand RNA viruses. Its viral RNA is translated to produce a polyprotein precursor that is further processed into three structural and seven non-structural proteins. The non-structural protein 3 (NS3) possess both protease and helicase activities. The C-terminal portion of the NS3 contains the ATPase/helicase domain presumably involved in viral replication. This domain has been expressed in Escherichia coli , purified in soluble form and structurally characterized. As judged by analytical centrifugation and size exclusion chromatography, the purified enzyme behaves as a monomer in solution. It has ATPase activity that is stimulated by the presence of RNA and single-stranded DNA molecules (ssDNA). However, we were unable to detect helicase activity at protein concentrations up to 500 nM. It has been reported that longer constructions of NS3 helicase domains from other flavivirus, like those which include residues of the linker region between the protease and the helicase domains, have helicase activity. Since all the conformational features of the purified WNV NS3 domain are those of a native protein, it is tempting to assume that the linker region plays a critical role in determining the protein–protein interactions that leads to the formation of the active oligomer.