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

作者：Fabritz Sebastian; Maass Franziska; Avrutina Olga; Heiseler Tim; Steinmann Bjoern; Kolmar Harald^*

来源：AMB Express, 2012, 2: 51.

DOI：10.1186/2191-0855-2-51

• Alcohol oxidase
• Haloalkane dehalogenase
• Haloalkanes
• Horseradish peroxidase
• Multistage enzyme reaction

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.

• 出版日期2012