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. 1999 Nov 15;344(Pt 1):273–280. doi: 10.1042/0264-6021:3440273

Cloning and molecular characterization of a soluble epoxide hydrolase from Aspergillus niger that is related to mammalian microsomal epoxide hydrolase.

M Arand 1, H Hemmer 1, H Dürk 1, J Baratti 1, A Archelas 1, R Furstoss 1, F Oesch 1
PMCID: PMC1220641  PMID: 10548561

Abstract

Aspergillus niger strain LCP521 harbours a highly processive epoxide hydrolase (EH) that is of particular interest for the enantioselective bio-organic synthesis of fine chemicals. In the present work, we report the isolation of the gene and cDNA for this EH by use of inverse PCR. The gene is composed of nine exons, the first of which is apparently non-coding. The deduced protein of the A. niger EH shares significant sequence similarity with the mammalian microsomal EHs (mEH). In contrast to these, however, the protein from A. niger lacks the common N-terminal membrane anchor, in line with the fact that this enzyme is, indeed, soluble in its native environment. Recombinant expression of the isolated cDNA in Escherichia coli yielded a fully active EH with similar characteristics to the fungal enzyme. Sequence comparison with mammalian EHs suggested that Asp(192), Asp(348) and His(374) constituted the catalytic triad of the fungal EH. This was subsequently substantiated by the analysis of respective mutants constructed by site-directed mutagenesis. The presence of an aspartic acid residue in the charge-relay system of the A. niger enzyme, in contrast to a glutamic acid residue in the respective position of all mEHs analysed to date, may be one important contributor to the exceptionally high turnover number of the fungal enzyme when compared with its mammalian relatives. Recombinant expression of the enzyme in E. coli offers a versatile tool for the bio-organic chemist for the chiral synthesis of a variety of fine chemicals.

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Selected References

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