• Journal of Life Science
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• v.12 no.5
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• pp.584-588
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• 2002

Asymmetric enantioselective resolution system using epoxide hydrolase activity of Aspergillus niger LK was developed and operated for the production of optically pure styrene oxide. Two-phase hollow-fiber reactor system was employed for the enhanced solubility of racemic styrene oxide in organic phase and protection of epoxide hydrolase activity in aqueous phase. For the removal of phenyl-1,2-ethandiol, the inhibitor of epoxide hydrolase, cascade hollow-fiber reactor system was also developed. Chiral (S)-styrene oxide (39 mM in dodecane) could be asymmetrically resolved with high enantiopurity (> 99% ee) using these reactor system.

• Journal of Institute of Control, Robotics and Systems
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• v.11 no.10
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• pp.857-863
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• 2005

A yeast strain utilizing styrene epoxide as a sole carbon and energy source was isolated from soil samples for the production of enantiopure of styrene epoxide by kinetic resolution. The strain, identified as a Rhodosporidium toruloides SJ-4, expressed an epoxide hydrolase which preferentially converted (R)-styrene epoxide into the corresponding diol. A maximum activity of 135 U/L was observed when biomass (dry cell mass) reached 6.7 g/L at 21 h of batch culture. Under the partially optimized reaction conditions ($35^{\circ}C$ and pH 8.0), the optically pure (S)-styrene epoxide was obtained with the yield of 21% when the initial substrate concentration was 100 mM. The reaction was completed at 9 h.

• KSBB Journal
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• v.23 no.5
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• pp.369-374
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• 2008

The effects of reaction temperature and the addition of various detergents on the enantioselective hyrolysis activity of the recombinant Escherichia coli containing the epoxide hydrolase (EH) gene of Rhodotorula glutinis were investigated for the production of enantiopure styrene oxide. The recombinant E. coli harboring the EH gene from R. glutinis exhibited the enantiopreference toward (R)-styrene oxide with the maximum hydrolytic activity of $165.04{\mu}mol/min/mg$ of dry cell weight (dcw). The addition of 0.5% (w/v) Tween 20 at $10^{\circ}C$ increased the initial hydrolysis rate and enantioselectivity by 1.45-fold and 2.0-fold, respectively. Enantiopure (S)-styrene oxide was prepared with 99% ee enantiopurity and 46.0% yield (theoretical yield=50%) from 20 mM racemic styrene oxide.

• Journal of Life Science
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• v.15 no.3 s.70
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• pp.332-336
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• 2005

Epoxide hydrolase activities of various microbial cells were analyzed by colorimetric assay based on alkylation of epoxides with 4-(p-nitrobenzyl)pyridine (NBP). The epoxide hydrolase activity was determined by measuring the decrease of color intensity at 560 nm due to the decrease of styrene oxide substrate by epoxide hydrolase-catalyzed hydrolysis reaction. The experimental conditions of NBP colorimetric assay were optimized for the efficient measurement of epoxide hydrolase activities from various microbial cells.

• Applied Chemistry for Engineering
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• v.18 no.3
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• pp.279-283
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• 2007

Epoxide hydrolase (EH) gene of Caulobacter crescentus was cloned by PCR and expressed in Escherichia coli. The C. crescentus EH (CcEH) primarily attacked at the benzylic carbon of (S)-styrene oxide, while the CcEH preferentially attacked at the terminal carbon of (R)-styrene oxide, thus leading to the formation of (R)-phenyl-1,2-ethanediol as the main product. (R)-phenyl-1,2-ethanediol was obtained with 85% enantiomeric excess and yield of 69% from racemic styrene oxide via enantioconvergent hydrolysis by using recombinant CcEH as the single biocatalyst.

• Applied Chemistry for Engineering
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• v.17 no.5
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• pp.557-560
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• 2006

The epoxide hydrolase (EH) of Aspergillus niger LK was expressed to high levels in Escherichia coli based on codon usage. E. coli, Rosetta (DE3)PLysS, containing a large number of tRNAs for rare-codons was employed as a host strain. The recombinant E. coli expressing A. niger EH showed an enhanced enantioselective hydrolysis activity toward racemic styrene oxide. Enantiopure (S)-styrene oxide with a high enantiopurity of 99% ee was obtained from racemic substrates.

• Journal of Life Science
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• v.16 no.3 s.76
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• pp.415-419
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• 2006

The epoxide hydrolase (EH) of Rhodotorula glutinis which has a high enantioselectivity against aromatic epoxide substrates was expressed to high levels in Escherichia coli based on codon usage. We analysed the Preference of codon usage between the yeast, R. glutinis, and bacteria, E. coli. E. coli, Rosetta(DE3)pLysS, harbors pRARE plasmid with tRNA genes for rare-codons was employed as a host strain. The recombinant E. coli expressing R. glutinis EH showed an enhanced enantioselective hydrolysis activity toward racemic styrene oxide. Enantiopure (S)-styrene oxide with a high enantiopurity of 99% ee (enantiomeric excess) was obtained from racemic substrates.

• Applied Chemistry for Engineering
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• v.19 no.5
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• pp.491-496
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• 2008

The microsomal epoxide hydrolase gene (referred to as mMCEH) of Mugil cephalus was cloned by PCR, and then inserted to pColdI and pET-21b(+) vector, respectively. The recombinant E. coli possessing the recombinant plasmids exhibited the enantioperference toward (R)-styrene oxide. When enantioselective kinetic resolutions were conducted with 20 mM racemic styrene oxide, enantiopure (S)-styrene oxide was obtained with high enantiopurity more than 99% enantiomeric excess (ee) and 24.50% yield by using the recombinant E. coli harboring pET-21b(+)/mMCEH.

• Microbiology and Biotechnology Letters
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• v.34 no.2
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• pp.129-135
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• 2006

A gene encoding for a putative microsomal epoxide hydrolase (mEH) of a zebrafish, Danio rerio, was cloned and characterized. The putative mEH protein of D. rerio exhibited sequence similarity with mammalian mEH and some other bacterial EHs. A structural model for the putative mEH was constructed using homology modeling based on the crystallographic templates, 1 qo7 and 1 ehy. The catalytic triad consisting of $Asp^{233}$, $Glu^{413}$, and $His^{440}$ was identified, and the characteristic features such as two tyrosine residues and oxyanion hole were found to be highly conserved. Based on bioinformatic analysis together with EH activity assay, the putative protein was annotated as mEH of D. rerio. Enantiopure styrene oxide with enantiopurity of 99%ee and yield of 33.5% was obtained from racemic styrene oxide by the enantioselective hydrolysis activity of recombinant mEH of D. rerio for 45 min.

• Applied Chemistry for Engineering
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• v.23 no.6
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• pp.599-603
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• 2012

The enantioselective hydrolysis of racemic styrene oxide in organic solvents was conducted using a recombinant E. coli expressing protein-engineered Mugil cephalus epoxide hydrolase (McEH). The volumetric total activity of the recombinant E. coli was enhanced 2.2-fold by IPTG induction at a mid-exponential growth phase. Among organic solvents with different log P values, isooctane was chosen based on the high activity and the enantioselectivity of McEH. Some lyoprotectants such as skim milk or sucrose enhanced the McEH activity. Enantiopure (S)-Styrene oxide with a 98% ee was obtained from the racemic styrene oxide with a 53.6% yield based on its theoretical yield in isooctane.