Cloning, expression and characterization of enantioselective epoxide hydrolases from marine microorganisms

Abstract

Epoxide hydrolases are ubiquitously found, catalyzing the conversion of epoxides to their corresponding vicinal diols. Previously we reported that 10 strains belonging to Erythrobacter showed epoxide hydrolase (EHase) activities toward various epoxide substrates. Three genes encoding putative EHases were identified by analyzing open reading frames (ORFs) of E. litoralis HTCC2594. Despite low similarities to reported EHases, the phylogenetic analysis of the three genes showed that eeh1 was similar to microsomal EHase while eeh2 and eeh3 could be grouped with soluble EHases. The three EHase genes were cloned, and the recombinant proteins (rEEH1, rEEH2 and rEEH3) were purified. The functionality of purified proteins was proved by hydrolytic activities toward styrene oxide. EEH1 preferentially hydrolyzed (R)-styrene oxide whereas EEH3 preferred to hydrolyze (S)-styrene oxide, representing enatioselective hydrolysis of styrene oxide. On the other hand, EEH2 could hydrolyze (R)- and (S)-styrene oxide at an equal rate. The optimal pH and temperature for the EHases occurred largely at neutal pHs and 40-55 oC. This is the first representation that a strict marine microorganism possessed three EHases with different enantioselectivity toward styrene oxide.