Development of an autofluorescent Pseudomonas nitroreducens with dehydrochlorinase activity for efficient...

Biodegradation or bioremediation is a more efficient and environmental friendly method for detoxification of hexachlorocyclohexane (HCH) residues compared to physical and chemical methods. Here, we report the functional expression of dehydrochlorinase (LinA) and enhanced green fluorescent protein (EGFP) in Pseudomonas nitroreducens for efficient biodegradation of gamma-HCH. The broad-host-range plasmid pVAG33, harboring dehydrochlorinase gene (linA) and enhanced green fluorescent protein gene (egfp), was constructed and transformed into the soil-isolated 1,2,4-trichlorobenzene (1,2,4-TCB)-degrading strain P. nitroreducens J5-1. Functional expression of LinA and EGFP was confirmed in the recombinant strain by Western-blotting analysis and by determining their enzymatic activities and fluorescence intensity. The recombinant strain could rapidly degrade 10microg ml(-1) gamma-HCH in 28h determined by GC-ECD analysis. It could completely mineralize gamma-HCH via gamma-HCH through 1,2,4-TCB and 3,4,6-trichlorocatechol and eventually entered the TCA cycle as determined by GC-MS analysis. The engineered strain can be applied in the form of a biocatalyst in a bioreactor for rapid degradation of HCH and chlorobenzene residues. Meanwhile, it can be easily monitored on-line by fluorescence of EGFP for its activity and fate.