NS5 polymerase stimulates RNA duplex unwinding by the Orthoflavivirus NS3 helicase

-ATP concentrations are required at a time coincident when NS3 encounters the RNA substrate. In addition, we find that Orthoflavivirus NS3s require translocation beyond the displaced strand to completely unwind a dsRNA substrate. Last, we find that Orthoflavivirus NS5 stimulates the ability of NS3 to unwind dsRNA. These results suggest that functional differences exist between the flavivirus NS3 helicases and illuminate that Orthoflavivirus NS3s require a functional interaction with the NS5 protein for coordination of its activity, as it is believed these two proteins constitute the viral replicase.