Characterization of the genes encoding reductive dehalogenases in Desulfitobacterium frappieri
PCP-1
Abstract Text
In the past, pentachlorophenol (PCP) has been used worldwide as a wood-preserving agent. Consequently, this toxic chemical has polluted, and still is, several soil and groundwater. Desulfitobacterium frappieri strain PCP-1 is the first anaerobic microorganism isolated that can dechlorinate PCP to 3-chlorophenol. Other halogenated compounds can also be dehalogenated by this strain at the ortho, meta and para positions. This bacterium utilizes two distinct dehalogenase systems: one for ortho dechlorination and a second for meta and para dechlorinations. Genes encoding for both systems named crdA and rdA1 were cloned and sequenced. Northern blot experiments showed that the expression of crdA is constitutive whether strain PCP-1 is cultured in presence or absence of chlorophenols. Using reverse transcribed polymerase chain reaction (RT-PCR), we found that rdA1 is expressed only when strain PCP-1 grows in presence of 3,5 dichlorophenols. Four putative chlorophenol reductive dehalogenase genes (HafcprA1 to 4) were found in Desulfitobacterium hafniense DCB-2 genome. It is known that strains DCB-2 and PCP-1 belong to the same species. Three of the four HafcprA where amplified from strain PCP-1 genome. In fact, the gene that was not amplified is responsible of the dehalogenation of 3-chloro-4-hydroxyphenylacetate in strain DCB-2; strain PCP-1 does not dehalogenate this molecule. Digoxigenin-labeled probes were generated for each gene found in this project, and Southern hybridization experiments were achieved with seven other strains of Desulfitobacterium species. The results showed that these bacteria contained several of these genes, suggesting that they could use different enzymes to achieve the dehalogenation of several chlorinated compounds.
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