Study of the enzymatic selectivity for peptides acylation : prediction of the selectivity of the Candida antarctica lipase B through molecular modeling approach and research of new specific aminoacylases enzymes

• Main Author: Ferrari, Florent
• Other Authors: Laboratoire Réactions et Génie des Procédés (LRGP), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Université de Lorraine, Isabelle Chevalot, Catherine Humeau-Virot
• Format: Doctoral or Postdoctoral Thesis
• Language: French
• Published: HAL CCSD 2014
• Subjects: Enzymatic selectivity; Peptide acylation; Candida antarctica lipase B; Acylases; Molecular modeling; Heterologous expression; Sélectivité enzymatique; Acylation peptidique; Lipase B de Candida antarctica; Modélisation moléculaire; Expression hétérologue; [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition; [SPI.GPROC]Engineering Sciences [physics]/Chemical and Process Engineering; Antarc*; Antarctica
• Online Access: https://tel.archives-ouvertes.fr/tel-01614465; https://tel.archives-ouvertes.fr/tel-01614465/document; https://tel.archives-ouvertes.fr/tel-01614465/file/2014LORR0252.pdf

Peptides exhibit various beneficial effects such as antioxidant, anti-hypertensive, neuroprotective, antiviral or antimicrobial activities. However, their use can be limited by their short half-life and their low biological availability. One solution to overcome these drawbacks is the acylation of peptides with fatty acids. This reaction called acylation can be catalyzed using enzymes. To date, very few studies focus on enzymatic acylation of peptides and on finding new enzymes catalyzing this reaction. The objectives of this work were, in a first time, to understand the selectivity mechanisms of the lipase B of Candida antarctica for peptides acylation combining experimental and molecular modeling approaches. This study highlighted enzyme/substrate interactions involved in the enzymatic selectivity and a modelexplaining the chemo- and regio-selectivity of this enzyme for peptide acylation reactions was built. In a second time, a preliminary study was carried out in order to identify new aminoacylase enzymes produced in the culture supernatant of various species of Streptomyces. These enzymes are able to catalyze acylation of peptides in aqueous media. A partial purification method was set and a comparative study was performed on the selectivity of C. antarctica lipase Band that of the new aminoacylases discovered in the culture supernatant of Streptomyces ambofaciens ATCC 23877. These enzymes presented a selectivity different from C. antarctica lipase B allowing the acylation of the N-terminal amino group of amino acids or peptides. A partial description of the aminoacylase activity of the supernatant crude extract of S. ambofaciens was performed. In a third and final part, a comparison of sequences of aminoacylases from Streptomyces mobaraensis with the genome of S.s ambofaciens ATCC 23877 was performed in order to identify genetic sequences encoding the new discovered aminoacylases from S. ambofaciens ATCC 23877. Each identified gene was deleted to correlate it with the aminoacylase activity observed in the ...