Contribution of sub-components to inhibitor potency to EV71 3C^pro were investigated

P2 groups need to be oriented to bind at the S2 subsite

S3 and S4 interacting moieties were required to generate sufficient potency

Hydrophobicity in S4 interacting moiety is helpful for cellular uptake

The data provide the basis for designing a new generation of inhibitors to 3C^pro