Molecular modeling and experimentation in solid/gas reactor for understanding the selectivity of lipase B from Candida antarctica

• Main Author: Marton, Zsuzsanna
• Other Authors: LIttoral ENvironnement et Sociétés - UMR 7266 (LIENSs), Institut national des sciences de l'Univers (INSU - CNRS)-La Rochelle Université (ULR)-Centre National de la Recherche Scientifique (CNRS), Université de La Rochelle, Marianne Graber
• Format: Doctoral or Postdoctoral Thesis
• Language: French
• Published: HAL CCSD 2010
• Subjects: Candida antarctica lipase B; Enantioselectivity; Solid/gas reactor; Molecular modeling; Inverse gas chromatography; Alcoholysis; Thermodynamic activity; Lipase B de Candida antarctica; Énantiosélectivité; Réacteur solide/gaz; Modélisation moléculaire; Chromatographie gazeuse en phase inverse; Alcoolyse; Activité thermodynamique; [SDV.SA]Life Sciences [q-bio]/Agricultural sciences; Antarc*; Antarctica
• Online Access: https://theses.hal.science/tel-00541104; https://theses.hal.science/tel-00541104/document; https://theses.hal.science/tel-00541104/file/these_ZsMarton_v3.pdf

The aim of this thesis was to understand more precisely the structural and environmental parameters governing the enantioselectivity of lipase B from Candida antarctica (CALB), involved in the discrimination of chiral secondary alcohols enantiomers. These compounds are used in particular for the synthesis of enantiomerically pure pharmaceutical molecules. Initially a systematic study of the orientation of butan-2-ol and pentan-2-ol in the active site was performed by molecular modeling. The results suggest the existence of additional binding modes to those mentioned in the literature. The potential energy comparison of the most stable conformations of the substrate, combined with the existence of non productive binding modes, allowed us to explain qualitatively the enantiopreference of CALB for the R form. Using the solid/gas reactor, we have shown that hydrophobic residues forming the access channel of substrate to the active site (Ile189, Leu278 and Ala282), played a significant role in the enantioselectivity of CALB towards secondary alcohols. The influence of the alkoxy part of the acyl donor on the enatiomeric ratio E has also been highlighted. Furthermore, we showed that isosteric mutations of the stereoselective pocket led to changes in pentan-2-ol activation thermodynamic parameters of acylation, probably due to changes in the hydrogen bonds network formed between residues of the pocket. Studies of water and substrates adsorption on the immobilized enzyme allowed us to relate the hydration status of CALB with specific effects of water on the enzyme activity and selectivity. L’objectif de cette thèse était de mieux comprendre les paramètres structuraux et environnementaux gouvernant l’énantiosélectivité de la lipase B de Candida antarctica (CALB), lors de la résolution des alcools secondaires linéaires chiraux. Ces composés sont notamment utilisés comme synthons chiraux dans l’industrie pharmaceutique. Dans un premier temps, une étude systématique des orientations du butan-2-ol et du pentan-2-ol au sein ...