title: Rational design of esterase BioH with enhanced enantioselectivity towards methyl (S)-o-chloromandelate.
abstract: Methyl (R)-o-chloromandelate (R-CMM) is an intermediate for the platelet aggregation inhibitor clopidogrel. Its preparation through enzymatic resolution of the corresponding ester has been hindered by the lack of an enzyme with satisfying enantioselectivity and activity. In the present work, we aimed to improve the enzymatic enantioselectivity towards methyl (S)-o-chloromandelate (S-CMM) by rational design, using esterase BioH as a model enzyme. Based on the differences in the binding mode of S- and R-enantiomers at the active cavity of the enzyme, the steric and electronic interactions between the key amino acids of BioH and the enantiomers were finely tuned. The enantioselectivity of esterase BioH towards S-CMM was improved from 3.3 (the wild type) to 73.4 (L123V/L181A/L207F). Synergistic interaction was observed between point mutations, and insight into the source of enzymatic enantioselectivity was gained by molecular dynamics simulations. The results can provide a reference for the enzyme design of other enzymes towards S-CMM for the enhancement of enantioselectivity.
authors.affiliation: Institute of Bioengineering, Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Jiali
authors.initials: J
authors.lastname: Gu
authors.forename: Lidan
authors.initials: L
authors.lastname: Ye
authors.forename: Fei
authors.initials: F
authors.lastname: Guo
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Lv
authors.forename: Wenqiang
authors.initials: W
authors.lastname: Lu
authors.forename: Hongwei
authors.initials: H
authors.lastname: Yu
mesh_terms: D020134:Catalytic Domain; D004950:Esterases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008333:Mandelic Acids / Q000378:metabolism*; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011487:Protein Conformation; D015202:Protein Engineering; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D008333:Mandelic Acids; D050505:Mutant Proteins; C522562:methyl 2-chloromandelate; D004950:Esterases