Molecular Modeling of Quercetin Acetylation by Lipases: Study of Enzyme-Substrate Interactions

• Main Author: Bidouil, Christelle
• Other Authors: Laboratoire d'Ingénierie des Biomolécules (LIBio), Université de Lorraine (UL), Université de Lorraine, Jean-Marc Engasser, Catherine Humeau-Virot
• Format: Doctoral or Postdoctoral Thesis
• Language: French
• Published: HAL CCSD 2012
• Subjects: Quercetin; Candida antarctica Lipase B (CALB); Acetylation; Molecular modeling; Docking; Molecular Dynamics; Quercétine; Lipase B de Candida antarctica; Modélisation moléculaire; Dynamique moléculaire; Flavonoïdes; Lipases; [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition; Antarc*; Antarctica
• Online Access: https://hal.univ-lorraine.fr/tel-01749457; https://hal.univ-lorraine.fr/tel-01749457/document; https://hal.univ-lorraine.fr/tel-01749457/file/DDOC_T_2012_0263_BIDOUIL.pdf

Accès restreint aux membres de l'Université de Lorraine jusqu'au 2015-12-01 Quercetin (QCT) is a plant-produced polyphenolic compound well-known for its antioxidant activities and beneficial health effects. Its solubility, stability, bioavailability and biological activities may be improved by a selective acylation of its hydroxyl groups. This work aims at studying the possibility of QCT enzymatic acetylation by Candida antarctica lipase B (CALB), the most industrially exploited lipase for regio- and enantioselective esterifications. In prospect of the rational enzyme design, a molecular modeling approach was implemented to understand the interactions that govern the substrate positioning and orientation in the lipase's active site. In a first experimental part, the absence of CALB acetylation activity towards quercetin in excess of vinyl acetate was confirmed. In a second part, this inactivity of CALB was explained by means of docking and molecular dynamics simulations. This results from an inappropriate positioning of the acyl donor linked to the catalytic serine and from an insufficient proximity of QCT hydroxyls vis-à-vis catalytic residues. The distance of QCT from the catalytic triad is due to its rigidity and to the narrow active site as well as to hydrophobic and electrostatic interactions between the substrate and the cavity residues. On the contrary, this molecular simulation approach predicts an appropriate positioning of both substrates in the active site of Pseudomonas cepacia lipase (PCL), which can perform QCT acetylation. In a third part, the impact of mutations of two residues implicated in the stabilization of QCT by hydrophobic interactions in CALB was investigated through simulations. The substitution of isoleucines by alanines and valines led to an increase in the catalytic pocket volume which intensified the mobility of QCT. However, these mutations are insufficient to allow an appropriate positioning of acetate and QCT in relation to the catalytic triad. The last part of this work ...