Subsite specificity of trypanosomal cathepsin L-like cysteine proteases : Probing the S2 pocket with phenylalanine-derived amino acids

Lecaille Fabien, Authié Edith, Moreau Thierry, Serveau Carole, Lalmanach Gilles. 2001. Subsite specificity of trypanosomal cathepsin L-like cysteine proteases : Probing the S2 pocket with phenylalanine-derived amino acids. European Journal of Biochemistry, 268 (9) : pp. 2733-2741.

Journal article ; Article de revue à facteur d'impact
[img] Published version - Anglais
Access restricted to CIRAD agents
Use under authorization by the author or CIRAD.

Télécharger (667kB)

Abstract : The S2 subsite of mammalian cysteine proteinases of the papain family is essential for specificity. Among natural amino acids, all these enzymes prefer bulky hydrophobic residues such as phenylalanine at P2. This holds true for their trypanosomal counterparts: cruzain from Trypanosoma cruzi and congopain from T. congolense. A detailed analysis of the S2 specificity of parasitic proteases was performed to gain information that might he of interest for the design of more selective pseudopeptidyl inhibitors. Nonproteogenic phenylalanyl analogs (Xaa) have been introduced into position P2 of fluorogenic substrates dansyl-Xaa-Arg-Ala-Pro-Trp, and their kinetic constants (Km, Kcat/Km) have been determined with congopain and cruzain, and related host cathepsins B and L. Trypanosomal cysteine proteases are poorly stereoselective towards D/L-Phe, the inversion of chirality modifying the efficiency of the reaction but not the Km. Congopain binds cyclohexylalanine better than aromatic Phe derivatives. Another characteristic feature of congopain compared to cruzain and cathepsins B and L was that it could accomodate a phenylglycyl residue (kcat/Km = 1300 mM-1.s-1), while lengthening of the side chain by a methylene group only slightly impaired the specificity constant towards trypanosomal cysteine proteases. Mono- and di-halogenation or nitration of Phe did not affect Km for cathepsin L-like enzymes, but the presence of constrained Phe derivatives prevented a correct fitting into the S2 subsite. A model of congopain has been built to study the fit of Phe analogs within the S2 pocket. Phe analogs adopted a positioning within the S2 pocket similar to that of the Tyr of the cruzain/Z-Tyr-Ala-fluoromethylke-tone complex. However, cyclohexylalanine has an energetically favorable chair-like conformation and can penetrate deeper into the subsite. Fitting of modeled Phe analogs were in good agreement with kinetic parameters. Furthermore, a linear relationship could be established with logP, supporting the suggestion that fitting into the S2 pocket of trypanosomal cysteine proteases depends on the hydrophobicity of Phe analogs. (Résumé d'auteur)

Classification Agris : L73 - Animal diseases

Auteurs et affiliations

  • Lecaille Fabien, Université François Rabelais (FRA)
  • Moreau Thierry, Université François Rabelais (FRA)
  • Serveau Carole, Université François Rabelais (FRA)
  • Lalmanach Gilles, Université François Rabelais (FRA)

Autres liens de la publication

Source : Cirad - Agritrop (

View Item (staff only) View Item (staff only)

[ Page générée et mise en cache le 2020-10-27 ]