PPNDS inhibits murine Norovirus RNA-dependent RNA-polymerase mimicking two RNA stacking bases

•Norovirus (NV) is a major cause of gastroenteritis worldwide.•Antivirals aiming at controlling and preventing NV infection are on demand.•PPNDS has been characterized as NV-RdRp inhibitor.•The mNV-RdRp/PPNDS 3D structure supports the design of anti-NV drugs.•PPNDS mimics the stacking interaction of two RNA-bases. Norovirus (NV) is a major cause of gastroenteritis worldwide. Antivirals against such important pathogens are on demand. Among the viral proteins that orchestrate viral replication, RNA-dependent-RNA-polymerase (RdRp) is a promising drug development target. From an in silico-docking search focused on the RdRp active site, we selected the compound PPNDS, which showed low micromolar IC50vs. murine NV-RdRp in vitro. We report the crystal structure of the murine NV-RdRp/PPNDS complex showing that two molecules of the inhibitor bind in antiparallel stacking interaction, properly oriented to block exit of the newly synthesized RNA. Such inhibitor-binding mode mimics two stacked nucleotide-bases of the RdRp/ssRNA complex.