Mutations in the stereospecificity pocket and at the entrance of the active site of Candida antarctica lipase B enhancing enzyme enantioselectivity

• Main Authors: Marton, Zsuzsanna, Léonard, Valérie, Syren, Per-Olof, Bauer, Cédric, Lamare, Sylvain, Hult, Karl, Tran, Vinh, Graber, Marianne
• 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), Department of Biochemistry KTH, Royal Institute of Technology Stockholm (KTH ), Department of Biochemistry- KTH, UMR CNRS 6204 U3B, Unité de Biotechnologie, Biocatalyse et Biorégulation (U3B), Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)
• Format: Article in Journal/Newspaper
• Language: English
• Published: HAL CCSD 2010
• Subjects: [SDV.BIO]Life Sciences [q-bio]/Biotechnology; [INFO.INFO-BT]Computer Science [cs]/Biotechnology; Double Point; Antarc*; Antarctica
• Online Access: https://hal.science/hal-00647795; https://hal.science/hal-00647795/document; https://hal.science/hal-00647795/file/MOLCAB-D-09-00286_corrected.pdf

International audience Two different parts of Candida antarctica lipase B (stereospecificity pocket at the bottom of the active site and hydrophobic tunnel leading to the active site) were redesigned by single- or double-point mutations, in order to better control and improve enzyme enantioselectivity toward secondary alcohols. Single-point isosteric mutations of Ser47 and Thr42 situated in the stereospecificity pocket gave rise to variants with doubled enantioselectivity toward pentan-2-ol, in solid/gas reactor. Besides, the width and shape of the hydrophobic tunnel leading to the active site was modified by producing the following single-point mutants: Ile189Ala, Leu278Val and Ala282Leu. For each of these variants a significant modification of enantioselectivity was observed compared to wild-type enzyme, indicating that discrimination of the enantiomers by the enzyme could also arise from their different accessibilities from the enzyme surface to the catalytic site.