Fig. 2.

(A) Superposition of residues lining the P1 pocket of Influenza C HEF (carbon atoms, cyan), MHV-DVIM HE (carbon atoms, green), and RCoV-NJ HE (carbon atoms, salmon). Surface representation is that of MHV-DVIM HE. Conserved residues within the SGNH family of hydrolases are underlined. (B) Surface representation of the catalytic center of MHV-DVIM HE with the P1 and P2 pockets indicated. The Ser-His-Asp catalytic triad is shown as sticks. (C) 9-N-Ac-Sia binding in the HEF catalytic site as observed in the crystal complex (18). Contacting amino acid side chains are shown in stick representation and colored by atom type (oxygen, red; nitrogen, blue; carbons, gray or green for amino acid side chains and 9-N-Ac-Sia, respectively). Oxyanion hole hydrogen bonds and the bidentate hydrogen bond interaction between Arg³⁰⁵ and the Sia carboxylate moiety are shown as black, dashed lines. (D) Model of 9-N-Ac-Sia binding in the MHV-DVIM HE catalytic site based on superposition with the HEF–inhibitor complex (carbon atoms, green) and on automated molecular docking (carbon atoms, salmon), represented as in Fig. 2C. (E) Catalytic activity of MHV-DVIM HE toward glycosidically bound 9-O-Ac-Sia is abrogated by substitution of Arg³⁰⁵ by Ala. Receptor destruction was assessed as in Fig. 1B. (F) Arg³⁰⁵Ala substitution in MHV-DVIM HE does not affect activity toward the synthetic substrate pNPA. Ser⁴⁴Ala is a catalytically inactive mutant. Enzymatic activity shown as percentage of wild-type activity.