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AMB Express. 2012; 2: 51.
Published online Sep 24, 2012. 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
Sebastian Fabritz,#1 Franziska Maaß,#1 Olga Avrutina,1 Tim Heiseler,1 Björn Steinmann,1 and Harald Kolmarcorresponding author1
1Institute of Organic Chemistry and Biochemistry, Technische Universität Darmstadt, Petersenstrasse 22, D-64287, Darmstadt, Germany
corresponding authorCorresponding author.
#Contributed equally.
Sebastian Fabritz: Fabritz/at/Biochemie-TUD.de; Franziska Maaß: Maass/at/Biochemie-TUD.de; Olga Avrutina: Avrutina/at/Biochemie-TUD.de; Tim Heiseler: Heiseler/at/Biochemie-TUD.de; Björn Steinmann: Steinmann/at/Biochemie-TUD.de; Harald Kolmar: Kolmar/at/Biochemie-TUD.de
Received August 10, 2012; Accepted September 16, 2012.
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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