Structure-Guided Regulation in the Enantioselectivity of an Epoxide Hydrolase to Produce Enantiomeric Monosubstituted Epoxides and Vicinal Diols via Kinetic Resolution.

Structure-guided microtuning of an epoxide hydrolase was executed. One mutant, A214C/A250I, displayed a 12.6-fold enhanced enantiomeric ratio ( = 202) toward -styrene oxide, achieving its nearly perfect kinetic resolution at 0.

Structure-guided improvement in the enantioselectivity of an Aspergillus usamii epoxide hydrolase for the gram-scale kinetic resolution of ortho-trifluoromethyl styrene oxide.

Microtuning the substrate-binding pocket (SBP) of EHs has emerged as an effective approach to manipulate their enantio- or regio-selectivities and activities towards target substrates. Here, the enantioselectivity (enantiomeric ratio, E) of AuEH2 towards a racemic (rac-) ortho-trifluoromethyl styrene oxide (o-TFMSO) was improved via microtuning its SBP. Based on the analysis on the crystal structure of AuEH2, its specific residues I192, Y216, R322 and L344 lining the SBP in close to the catalytic triad were identified for site-saturation mutagenesis.

Nearly perfect kinetic resolution of racemic o-nitrostyrene oxide by AuEH2, a microsomal epoxide hydrolase from Aspergillus usamii, with high enantio- and regio-selectivity.

Only a few known epoxide hydrolases (EHs) displayed activity towards o-nitrostyrene oxide (4a), presumably owing to the large steric hindrance caused by o-nitro substituent. Therefore, excavating EHs with high activity and enantio- and/or regio-selectivity towards racemic (rac-) 4a is essential but challenging. Here, AuEH2 from Aspergillus usamii was expressed in E.

Near-perfect kinetic resolution of o-methylphenyl glycidyl ether by RpEH, a novel epoxide hydrolase from Rhodotorula paludigena JNU001 with high stereoselectivity.

In order to provide more alternative epoxide hydrolases for industrial production, a novel cDNA gene Rpeh-encoding epoxide hydrolase (RpEH) of Rhodotorula paludigena JNU001 identified by 26S rDNA sequence analysis was amplified by RT-PCR. The open-reading frame (ORF) of Rpeh was 1236 bp encoding RpEH of 411 amino acids and was heterologously expressed in Escherichia coli BL21(DE3). The substrate spectrum of expressed RpEH showed that the transformant E.

Greatly enhancing the enantioselectivity of PvEH2, a Phaseolus vulgaris epoxide hydrolase, towards racemic 1,2-epoxyhexane via replacing its partial cap-loop.

To achieve the kinetic resolution and enantioconvergent hydrolysis of rac-1,2-epoxyhexane, the E value of PvEH2 was enhanced by substituting its partial cap-loop. Based on the experimental results reported previously and computer-aided analysis, the flexible and variable cap-loop, especially its middle segment, was speculated to be related to the catalytic properties of PvEH2. In view of this, four PvEH2's hybrids, Pv2St, Pv2Pv1, Pv2Vr1 and Pv2Vr2, were designed by substituting the middle segment (EGMGSNLNTSMP) of a cap-loop in PvEH2 with the corresponding ones in StEH, PvEH1, VrEH1 and VrEH2, respectively.

Manipulating regioselectivity of an epoxide hydrolase for single enzymatic synthesis of (R)-1,2-diols from racemic epoxides.

Both the activity and regioselectivity of Phaseolus vulgaris epoxide hydrolase were remarkably improved via reshaping two substrate tunnels based on rational design. The elegant one-step enantioconvergent hydrolysis of seven rac-epoxides was achieved by single mutants, allowing green and efficient access to valuable (R)-1,2 diols with high eep (90.1-98.

Substantially improving the enantioconvergence of PvEH1, a Phaseolus vulgaris epoxide hydrolase, towards m-chlorostyrene oxide by laboratory evolution.

Background: Epoxide hydrolase can regioselectively catalyze the oxirane ring-opening hydrolysis of rac-epoxides producing the corresponding chiral diols. In our laboratory, a gene named pveh1 encoding an EH from Phaseolus vulgaris was cloned. Although the directed modification of PvEH1 was carried out, the mutant PvEH1 showed a limited degree of enantioconvergence towards racemic (rac-) m-chlorostyrene oxide (mCSO).

Near-perfect kinetic resolution of racemic p-chlorostyrene oxide by SlEH1, a novel epoxide hydrolase from Solanum lycopersicum with extremely high enantioselectivity.

An open reading frame of sleh1, a gene encoding for a novel epoxide hydrolase from Solanum lycopersicum (SlEH1), was amplified by RT-PCR and expressed in E. coli BL21(DE3). The substrate spectrum assay showed that E.

Multiple site-directed mutagenesis of a Phaseolus vulgaris epoxide hydrolase to improve its catalytic performance towards p-chlorostyrene oxide based on the computer-aided re-design.

To improve the activity and regioselectivity of a Phaseolus vulgaris epoxide hydrolase (PvEH3) towards p-chlorostyrene oxide (pCSO), the site-directed mutagenesis was conducted based on the computer-aided re-design. Firstly, seven single-site variants of a PvEH3-encoding gene (pveh3) were constructed as designed theoretically and expressed in E. coli BL21(DE3), respectively.