Identification and catalytic properties of new epoxide hydrolases from the genomic data of soil bacteria

• Published in: Enzyme and microbial technology 2020-09, Vol.139, p.109592, Article 109592
• Main Authors: Stojanovski, Gorjan, Dobrijevic, Dragana, Hailes, Helen C., Ward, John M.
• Format: Article
• Language: English
• Subjects: Bacteria - enzymology; Bacteria - genetics; Biocatalysis; Biotransformation; Data Mining; Epoxide hydrolase; Epoxide Hydrolases - genetics; Epoxide Hydrolases - metabolism; Epoxy Compounds - chemistry; Escherichia coli - genetics; Genome mining; Genome, Bacterial; Hydrolysis; Limonene epoxide hydrolase; Soil Microbiology; Stereoisomerism; Substrate Specificity
• ISSN: 0141-0229; 1879-0909; 1879-0909
• DOI: 10.1016/j.enzmictec.2020.109592

•Five new active epoxide hydrolases were found.•The limonene epoxide hydrolase from pQR1982 has a wide substrate scope.•Enzymes from pQR1982 and pQR1984 had opposite epoxide ring-opening stereoselectivities.•Enzymes from pQR1984 and pQR1990 de-symmetrised the meso-epoxide, cyclohexene oxide.•The ⍺/β EH from pQR1984 had comparable activity in the presence of 5–30% MeOH. Epoxide hydrolases (EHs) catalyse the conversion of epoxides into vicinal diols. These enzymes have extensive value in biocatalysis as they can generate enantiopure epoxides and diols which are important and versatile synthetic intermediates for the fine chemical and pharmaceutical industries. Despite these benefits, they have seen limited use in the bioindustry and novel EHs continue to be reported in the literature. We identified twenty-nine putative EHs within the genomes of soil bacteria. Eight of these EHs were explored in terms of their activity. Two limonene epoxide hydrolases (LEHs) and one ⍺/β EH were active on a model compound styrene oxide and its ring-substituted derivatives, with low to good percentage conversions of 18–86%. Further exploration of the substrate scope with enantiopure (R)-styrene oxide and (S)-styrene oxide, showed different epoxide ring opening regioselectivities. Two enzymes, expressed from plasmids pQR1984 and pQR1990 de-symmetrised the meso-epoxide cyclohexene oxide, forming the (R,R)-diol with high enantioselectivity. Two LEHs, from plasmids pQR1980 and pQR1982 catalysed the hydrolysis of (+) and (−) limonene oxide, with diastereomeric preference for the (1S,2S,4R)- and (1R,2R,4S)-diol products, respectively. The enzyme from plasmid pQR1982 had a good substrate scope for a LEH, being active towards styrene oxide, its analogues, cyclohexene oxide and 1,2-epoxyhexane in addition to (±)-limonene oxide. The enzymes from plasmids pQR1982 and pQR1984 had good substrate scopes and their enzymatic properties were characterised with respect to styrene oxide. They had comparable temperature optima and pQR1984 had 70% activity in the presence of 40% of the green solvent MeOH, a useful property for bio-industrial applications. Overall, this study has provided novel EHs with potential value in industrial biocatalysis.