Cloning, expression and enantioselective hydrolytic catalysis of a microsomal epoxide hydrolase from a marine fish, Mugil cephalus

Abstract

The cDNA of a marine fish microsomal epoxide hydrolase (mEH) gene from Mugil cephalus was cloned by rapid amplification of cDNA ends (RACE) techniques. The homology model for the mEH of M. cephalus showed a characteristic structure of alpha/beta-hydrolase-fold main domain with a lid domain over the active site. The characteristic catalytic triad, consisting of Asp(238), His(444), and Glu(417), was highly conserved. The cloned mEH gene was expressed in Escherichia coli and the recombinant mEH exhibited (R)-preferred hydrolysis activity toward racemic styrene oxide. We obtained enantiopure (S)-styrene oxide with a high enantiopurity of more than 99% enantiomeric excess and yield of 15.4% by batch kinetic resolution of 20 mM racemic styrene oxide.

Functional expression and magnetic nanoparticle-based Immobilization of a protein-engineered marine fish epoxide hydrolase of Mugil cephalus for enantioselective hydrolysis of racemic styrene oxide

“…inverted triangle 100 mM Tris/HCl, open triangle 100 mM glycine/NaOH. For determination of the optimal reaction temperature, EH activities were measured at pH 7.4 over the different temperatures ranging from 10 to 50°C as described previously (Lee et al 2007) …”

“…They can be prepared by resolution of racemic epoxides by using epoxide hydrolase (EH) (Steinreiber and Faber 2001;Widersten et al 2010). Recently, a fish microsomal EH (mEH) gene from Mugil cephalus has been cloned and characterized (Lee et al 2007). The catalytic activity of M. cephalus EH has been enhanced 35-fold by sitedirected mutagenesis based on comparative modeling (Choi et al 2009).…”

Enantioselective Hydrolysis of Styrene Oxide and Benzyl Glycidyl Ether by a Variant of Epoxide Hydrolase from Agromyces mediolanus

“…Although kinetic resolution of racemic SO by EH has been reported in many papers, there are only a few EHs that have been obtained from marine resources. EH from the marine fish ( Mugil cephalus ) exhibited ( R )-preferred hydrolysis activity towards racemic SO, and ( S )-SO was obtained with >99% ee and yield of 15.4% [8]. The kinetic resolution of racemic SO also was performed by EH from a marine microorganism ( Erythrobacter spp), ( S )-SO with an ee > 99%, but only an 8.5% yield was obtained [14].…”

“…One alternative method for preparing chiral epoxides is the enantioselective hydrolysis of racemic epoxides using epoxide hydrolase (EH, EC 3.3.2.3), which have been discovered in and cloned from many organisms such as plants, mammals, insects, bacteria, fungi, and yeasts [6,7,8]. To date, several EHs have been used to prepare chiral SO, but the enantiomeric excess (ee) or yields are always low.…”

One-pot biotransformation of racemic styrene oxide into (R)-1,2-phenylethandiol by two recombinant microbial epoxide hydrolases

“…A novel marine fish EH gene from Mugil cephalus has been also cloned and characterized [12]. Recently, we have explored the enantioconvergent hydrolysis of racemic styrene oxide based on the enantio-flexibility of the C. crescentus EH (CCEH) and M. cephalus EH (MCEH) [13,14].…”