Transcriptional Regulation of the cpr Gene Cluster in ortho -Chlorophenol-Respiring Desulfitobacterium dehalogenans

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Abstract
To characterize the expression and possible regulation of reductive dehalogenation in halorespiring bacteria, a 11.5-kb genomic fragment containing the o-chlorophenol reductive dehalogenase-encoding cprBA genes of the gram-positive bacterium Desulfitobacterium dehalogenans was subjected to detailed molecular characterization. Sequence analysis revealed the presence of eight designated genes with the ordercprTKZEBACD and with the same polarity except forcprT. The deduced cprC and cprKgene products belong to the NirI/NosR and CRP-FNR families of transcription regulatory proteins, respectively. CprD and CprE are predicted to be molecular chaperones of the GroEL type, whereascprT may encode a homologue of the trigger factor folding catalysts. Northern blot analysis, reverse transcriptase PCR, and primer extension analysis were used to elucidate the transcriptional organization and regulation of the cpr gene cluster. Results indicated halorespiration-specific transcriptional induction of the monocistronic cprT gene and the biscistroniccprBA and cprZE genes. Occasional read-through at cprC gives rise to a tetracistronic cprBACDtranscript. Transcription of cprBA was induced 15-fold upon addition of the o-chlorophenolic substrate 3-chloro-4-hydroxyphenylacetic acid within 30 min with concomitant induction of dehalogenation activity. Putative regulatory protein binding motifs that to some extent resemble the FNR box were identified in the cprT-cprK and cprK-cprZ intergenic regions and the promoter at cprB, suggesting a role for FNR-like CprK in the control of expression of thecprTKZEBACD genes.