Structural Analysis of Inhibitor Binding to Enterovirus-D68 3C Protease

Figures

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  Figure 1. The conservation of amino acid sequences for 3C protease and viral polyprotein cleavage sites: (A) cleavage site sequences in EV68 strains and (B) 3C protease sequences across picornaviruses, highlighting the conserved regions with darker shades.

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  Figure 2. Co-crystal structures of EV68-3C protease with inhibitors: (A) irreversible covalent inhibitors rupintrivir (slate blue) and (B) AG7404 (green) and (C) reversible covalent inhibitor GC-376 (pink). On the left panels, the protease is depicted as a gray surface while the inhibitor is represented as a colored stick within the electron density (2F0-FC map) and hydrogen bond interactions (yellow dashed lines) shown with the surrounding protease residues (gray sticks). The catalytic dyad residues (H40 and C147) are highlighted in yellow. The chemical structures of the bound inhibitors and hydrogen bonds with the EV68-3C protease residues are displayed on the right.

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  Figure 3. Coordination of an ordered water molecule within the S1 pocket of EV68-3C protease in ligand-bound crystal structures: (A) 3B3C peptide (purple sticks) PDB IDs: 9AX9 and (B–D) inhibitors (GC-376 in pink, rupintrivir in slate blue, and AG7404 in green, respectively) bound within EV68-3C protease. The hydrogen bonds between the water (red sphere) and surrounding EV68-3C protease residues (Thr142, Asn165, and Gly166) are depicted as dashed lines.

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  Figure 4. Fit of inhibitors within the EV68-3C protease substrate envelope: inhibitors (A) AG7404 (green sticks) and (B) GC-376 (pink sticks). The protease is shown as a gray surface with residues around the inhibitor represented as sticks (gray) and labeled. The inhibitor from the co-crystal structures has been superimposed onto the substrate envelope (cyan volume). Red arrows highlight regions of the inhibitor protruding from the substrate envelope.