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Applied and Environmental Microbiology, April 2006, p. 2905-2917, Vol. 72, No. 4
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.4.2905-2917.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Diversity and Biocatalytic Potential of Epoxide Hydrolases Identified by Genome Analysis{dagger}

Bert van Loo,1,{ddagger} Jaap Kingma,1 Michael Arand,2 Marcel G. Wubbolts,3 and Dick B. Janssen1*

Biochemical Laboratory, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands,1 Institute of Pharmacology and Toxicology, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland,2 DSM Pharma Chemicals, Advanced Synthesis, Catalysis & Development, DSM Research, P.O. Box 18, 6160 MD Geleen, The Netherlands3

Received 5 September 2005/ Accepted 19 January 2006

Epoxide hydrolases play an important role in the biodegradation of organic compounds and are potentially useful in enantioselective biocatalysis. An analysis of various genomic databases revealed that about 20% of sequenced organisms contain one or more putative epoxide hydrolase genes. They were found in all domains of life, and many fungi and actinobacteria contain several putative epoxide hydrolase-encoding genes. Multiple sequence alignments of epoxide hydrolases with other known and putative {alpha}/ß-hydrolase fold enzymes that possess a nucleophilic aspartate revealed that these enzymes can be classified into eight phylogenetic groups that all contain putative epoxide hydrolases. To determine their catalytic activities, 10 putative bacterial epoxide hydrolase genes and 2 known bacterial epoxide hydrolase genes were cloned and overexpressed in Escherichia coli. The production of active enzyme was strongly improved by fusion to the maltose binding protein (MalE), which prevented inclusion body formation and facilitated protein purification. Eight of the 12 fusion proteins were active toward one or more of the 21 epoxides that were tested, and they converted both terminal and nonterminal epoxides. Four of the new epoxide hydrolases showed an uncommon enantiopreference for meso-epoxides and/or terminal aromatic epoxides, which made them suitable for the production of enantiopure (S,S)-diols and (R)-epoxides. The results show that the expression of epoxide hydrolase genes that are detected by analyses of genomic databases is a useful strategy for obtaining new biocatalysts.

* Corresponding author. Mailing address: Biochemical Laboratory, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands. Phone: 31 (0)50 3634008. Fax: 31 (0)50 3634165. E-mail: D.B.Janssen{at}rug.nl.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

{ddagger} Present address: Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge, United Kingdom.

Applied and Environmental Microbiology, April 2006, p. 2905-2917, Vol. 72, No. 4
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.4.2905-2917.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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