Regulation of anaerobic dehalorespiration by the transcriptional activator CprK.

The Journal of Biological Chemistry
Stelian M PopStephen W Ragsdale

Abstract

Desulfomonile, Desulfitobacterium, and Dehalobacter are anaerobic microbes that can derive energy from the reductive dehalogenation of chlorinated organic compounds, many of which are environmental pollutants. There is very little information about how anaerobic dehalorespiration is regulated. An open reading frame within the Desulfitobacterium dehalogenans chlorophenol reductase (cpr) gene cluster (cprK) was proposed to be a transcriptional regulatory protein (Smidt, H., van Leest, M., van der Oost, J., and deVos, W. M. (2000) J. Bacteriol. 182, 5683-5691). We have cloned, actively overexpressed in Escherichia coli, and purified to homogeneity the D. dehalogenans CprK. The results of electrophoretic mobility shift assays, DNA footprinting studies, and promoter-lac fusion experiments indicate that CprK is a transcriptional activator of the cpr gene cluster. CprK binds 3-chloro-4-hydroxyphenylacetate (CHPA) with high affinity (K(d) = 3.5 mum, determined by isothermal titration calorimetry), which promotes its specific interaction with a DNA sequence (TTAAT-N4-ACTAA) located upstream of the -35 and -10 promoter regions of several cpr genes and activates transcription of these genes. Binding to the upstream "box" sequence increase...Continue Reading

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