Premium
Probing the Mechanism of CAL‐B‐Catalyzed aza‐Michael Addition of Aniline Compounds with Acrylates Using Mutation and Molecular Docking Simulations
Author(s) -
Gu Bo,
E Hu Zu−,
Yang ZengJie,
Li Jun,
Zhou ZiWen,
Wang Na,
Yu XiaoQi
Publication year - 2019
Publication title -
chemistryselect
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.437
H-Index - 34
ISSN - 2365-6549
DOI - 10.1002/slct.201900112
Subject(s) - docking (animal) , active site , chemistry , stereochemistry , catalysis , aniline , stereocenter , candida antarctica , mutant , lipase , enzyme , combinatorial chemistry , organic chemistry , biochemistry , enantioselective synthesis , medicine , nursing , gene
CAL−B (Lipase B from Candida antarctica ) catalyzed aza‐Michael addition of a set of aniline compounds with acrylates under mild conditions was described. A systematic study allowed to determine the appropriate solvent, enzyme loading, reaction temperature and time. In order to speculate and verify its mechanism, the wild‐type and three mutants (S105 A, H224 A and I189 A) of CAL−B were expressed. Some control experiments demonstrated the active site was responsible for the enzymatic process, in which Ser105 and His224 played a crucial role. Besides, the mutation of Ile189 also affected its activity a lot. Based on these results, a docking experiment was performed to speculate the mechanism: the oxyanion hole (Thr40 and Gln106) of the active site activated the acrylates and stabilized the transition states. The Ile189 residues, as an important part of active cavity, could form a strong hydrophobic interaction with substrates. And the Ser105 and His224 residues were responsible for proton transfer during the catalytic process. This would help to understand the promiscuity of CAL−B, and provide ideas to design novel enzyme to improve the efficiency of its promiscuity.