Heterologous overexpression, purification and characterisation of an alcohol dehydrogenase (ADH2) from Halobacterium sp. NRC-1

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Title: Heterologous overexpression, purification and characterisation of an alcohol dehydrogenase (ADH2) from Halobacterium sp. NRC-1
Authors: Liliensiek, Ann-Kathrin
Cassidy, Jennifer
Gucciardo, Gabriele
Whitely, Cliadhna
Paradisi, Francesca
Permanent link: http://hdl.handle.net/10197/5422
Date: Oct-2013
Abstract: Replacement of chemical steps with biocatalytic ones is becoming increasingly more interesting due to the remarkable catalytic properties of enzymes, such as their wide range of substrate specificities and variety of chemo-, stereo- and regioselective reactions. This study presents characterization of an alcohol dehydrogenase (ADH) from the halophilic archaeum Halobacterium sp. NRC-1 (HsADH2). A hexahistidine-tagged recombinant version of HsADH2 (His-HsADH2) was heterologously overexpressed in Haloferax volcanii. The enzyme was purified in one step by immobilised Ni-affinity chromatography (IMAC). His-HsADH2 was halophilic and mildly thermophilic with optimal activity for ethanol oxidation at 4 M KCl around 60 °C and pH 10.0. The enzyme was extremely stable, retaining 80 % activity after 30 days. His-HsADH2 showed preference for NADP(H) but interestingly retained 60 % activity towards NADH. The enzyme displayed broad substrate specificity, with maximum activity obtained for 1-propanol. The enzyme also accepted secondary alcohols such as 2-butanol and even 1-phenylethanol. In the reductive reaction, working conditions for His-HsADH2 were optimised for acetaldehyde and found to be 4 M KCl and pH 6.0. His-HsADH2 displayed intrinsic organic solvent tolerance, which is highly relevant for biotechnological applications.
Funding Details: Science Foundation Ireland
Type of material: Journal Article
Publisher: Springer
Copyright (published version): 2013, Springer
Keywords: Halobacterium sp. NRC-1Haloferax volcaniiAlcohol dehydrogenaseSolvent tolerance
DOI: 10.1007/s12033-013-9666-4
Language: en
Status of Item: Peer reviewed
Appears in Collections:Chemistry Research Collection

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