Fig. 5: Serial viral passage experiments to identify drug-resistant mutants against EV-D68 2C inhibitor Jun6504 and MD simulations of Jun6504 with EV-D68 2C WT and D183V mutant.
From: A rationally designed 2C inhibitor prevents enterovirus D68-infected mice from developing paralysis
a Schematic illustration of the serial passage of Jun6504 in RD cells with EV-D68 US/MO/14-18947. The graph was created with Biorender.com. Li, K (2025) https://BioRender.com/0kxixma. b Electropherogram traces of 2C protein coding region at residues 112, 183, and 323 from viruses at passages 0, 3, 6, and 9. c Antiviral EC50 curves of Jun6504 against recombinant EV-D68 viruses in plaque assay. The EC50 values are mean ± SD from two biological replicates. d Thermal shift binding assay of Jun6504 with EV-D68 2C WT, D183V, D323G, and D183V/D323G. The red line indicates a 0.5 °C shift (ΔTm = 0.5 °C). A ΔTm greater than 0.5 °C generally indicates ligand binding to the protein69. The assay was performed in two biological replicates. e Interactions between Jun6504 and the residues at the allosteric site of EV-D68 2C. f Stabilizing interactions inside the binding area of 2C with Jun6504; hydrogen bonding interactions bar is depicted in green, ion hydrogen bonding interactions in red, water bridges in blue, and lipophilic interactions in gray. g RMSD plots of Cα carbons of 2C (blue diagram) and of the heavy atoms of the ligand Jun6504 (red diagram). h Interactions between Jun6504 and the residues at the allosteric site of EV-D68 2C-D183V. i Stabilizing interactions inside the binding area of 2C-D183V with Jun6504; hydrogen bonding interactions bar is depicted in green, ion hydrogen bonding interactions in red, water bridges in blue and lipophilic interactions in gray. j RMSD plots of Cα carbons of 2C-D183V (blue diagram) and of the heavy atoms of the ligand Jun6504 (red diagram). Source data are provided as a Source Data file.
