(A) Structure of EV71 protomer in complex with NLD. Icosahedral protomeric unit of EV71 (PDBid 4CEY), viewed from the inside of the capsid. VP1, blue, VP2, green, VP3 red, VP4 yellow, NLD magenta. Protein shown in cartoon representation, NLD as spheres, with the chemical structures of GPP3, NLD and ALD molecules reported below. (B) Structure of EV71 pocket in complex with NLD. EV71 (PDBid 4CEY) showing a close-up of the VP1 (blue) pocket with NLD (magenta) in-place. The side-chains of F135, F155, D112 and I113 and NLD are shown as ball-and-sticks. Hydrogen bonds are shown as dashed (C) Predicted structure of inhibitor resistant mutant in complex with NLD. EV71 (PDBid 4CEY) in blue, with VP1 model generated by Rossetta shown in grey. The two mutated side chains I113M and V123I are shown as grey balls-and-sticks. The distance between the M113 side chain and NLD linker, corresponding to 2.1 Å, is shown as a dotted line. (D) Structure of CVA16 (PDBid:5ABJ) in complex with GPP3. Close-up view of the VP1 pocket (in blue) with GPP3 bound (magenta). The side chains of Y135 and F155 are shown as blue ball-and-sticks. The predicted structure of the inhibitor-resistant mutant in complex with GPP3 is shown in grey with the L113F mutation in grey ball-and-sticks. In all ball-and-stick representation, nitrogen atoms are blue, oxygen red and sulphur yellow. Sequencing shows a change to M or F could be achieved by a single point mutation for both EV71 and CVA16. Comparison with the NCBI gene bank database revealed that none of the 5830 EV71 or 248 CVA16 sequences deposited contained any combination of these mutations.