Fig. 5

Structural representation of the FhCL3 zymogen highlighting the interaction of the propeptide with the mature enzyme. (a) The 3-D modelling of FhCL3 zymogen suggests the importance of key ppFhCL3 amino acid residues (pink) that stabilize the propeptide binding/inhibition (in pink) with the FhCL3 mature domain (grey; residues and labels in cyan and black). The ppFhCL3 residue Tyr⁴⁶ binds to Asp²²⁸ and Phe²³⁷, while Lys⁴⁷ binds to Ser²³⁴ and Glu²³³ residues within the propeptide binding loop (PBL), respectively. The C-terminal portion of the propeptide is then inserted into the substrate binding cleft, and specific interaction between the propeptide residue Leu⁶⁶ with the hydrophobic pocket of Ala²²⁵, Phe²²⁹, His²⁴⁹ and Trp²⁷¹, and Glu⁶⁸ with Thr²⁴⁸ in the mature enzyme result in a tight binding that prevent substrate entrance in the enzyme cleft. Dotted lines represent hydrogen bonds (black), a salt bridge (pink) and π-π interaction (cyan) formed between residues of the propeptide and the mature enzyme. (b) Primary amino acid sequence of the ppFhCL3. Residues highlighted in black are predicted to be involved in fundamental interactions with the mature enzyme domain to stabilize the binding. The sequence portion of the ppFhCL3 used as a template to produce a synthetic peptide (33 residues) is underlined. Residues highlighted in grey form the propeptide conserved motifs ERFNIN and GNFD