Targeting flavivirus RNA dependent RNA polymerase through a pyridobenzothiazole inhibitor

- From in silico docking we identified HeE1-2Tyr as inhibitor of flavivirus RdRp.
- The crystal structure of DENV3 RdRp with HeE1-2Tyr showed a possible interference of the inhibitor with template RNA.
- In contrast, kinetic data showed a non competitive inhibition mechanism.
- Several point mutations suggest an alternative binding site, hidden by the priming loop in crystal structure.

RNA dependent RNA polymerases (RdRp) are essential enzymes for flavivirus replication. Starting from an in silico docking analysis we identified a pyridobenzothiazole compound, HeE1-2Tyr, able to inhibit West Nile and Dengue RdRps activity in vitro, which proved effective against different flaviviruses in cell culture. Crystallographic data show that HeE1-2Tyr binds between the fingers domain and the priming loop of Dengue virus RdRp (Site 1). Conversely, enzyme kinetics, binding studies and mutational analyses suggest that, during the catalytic cycle and assembly of the RdRp-RNA complex, HeE1-2Tyr might be hosted in a distinct binding site (Site 2). RdRp mutational studies, driven by in silico docking analysis, allowed us to locate the inhibition Site 2 in the thumb domain. Taken together, our results provide innovative concepts for optimization of a new class of anti-flavivirus compounds.