Adenosine triphosphate analogs can efficiently inhibit the Zika virus RNA-dependent RNA polymerase.

We describe the expression and purification of an active recombinant Zika virus RNA-dependent RNA polymerase (RdRp). Next, we present the development and optimization of an in vitro assay to measure its activity. We then applied the assay to selected triphosphate analogs and discovered that 2'-C-methylated nucleosides exhibit strong inhibitory activity. Surprisingly, also carbocyclic derivatives with the carbohydrate locked in a North-like conformation as well as a ribonucleotide with a South conformation exhibited strong activity. Our results suggest that the traditional 2'-C-methylated nucleosides pursued in the race for anti-HCV treatment can be superseded by brand new scaffolds in the case of the Zika virus.