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AMB Express. 2012; 2: 51.
Published online 2012 September 24. doi:  10.1186/2191-0855-2-51
PMCID: PMC3487978

A sensitive method for rapid detection of alkyl halides and dehalogenase activity using a multistep enzyme assay

Abstract

A method for the detection of haloalkane conversion to the corresponding alcohols by haloalkane dehalogenases is described. It is based on a multistage enzyme reaction which allows for the analysis of alkyl halides in buffered systems. Irreversible hydrolytic dehalogenation catalyzed by haloalkane dehalogenase DhaA from Rhodococcus erythropolis transfers an alkyl halide into a corresponding alcohol that is further oxidized by alcohol oxidase AOX from Pichia pastoris yielding a respective aldehyde and hydrogen peroxide easily detectable via the horseradish peroxidase catalyzed oxidation of chromogenic molecules. Due to its high sensitivity (0.025 mM, 0.43 ppm for 1,3-dibromopropane), low expenditure and the ability of handling a large number of samples in parallel, this method is an attractive alternative to existing procedures for the monitoring of both haloalkanes and dehalogenases.

Keywords: Alcohol oxidase, Haloalkane dehalogenase, Haloalkanes, Horseradish peroxidase, Multistage enzyme reaction

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