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In vitro and in vivo characterization of three major dadAX promoters in Escherichia coli that are regulated by cyclic AMP-CRP and Lrp

Abstract

We have shown previously that the dadAX operon of Escherichia coli expresses multiple transcripts, which are repressed by leucine-responsive regulatory protein (Lrp). Here we used site-directed mutagenesis and in vitro and in vivo transcription assays to show that each of the three major dad transcripts requires a specific promoter. These promoters, P1-P3, overlap and are positively regulated in vivo by cyclic AMP-CRP. DNase I footprinting experiments localized two CRP binding sites in this region: CRP1, which is positioned upstream of P1-P3, and CRP2, which is located within the promoters. Site-directed mutagenesis of each site provided evidence that CRP1 is necessary for the effects of cyclic AMP-CRP on dad expression in vivo and in vitro, and that CRP2 probably plays little or no role in this process.

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Received: 14 August 1997 / Accepted: 19 February 1998

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Zhi, J., Mathew, E. & Freundlich, M. In vitro and in vivo characterization of three major dadAX promoters in Escherichia coli that are regulated by cyclic AMP-CRP and Lrp. Mol Gen Genet 258, 442–447 (1998). https://doi.org/10.1007/s004380050754

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  • Key wordsdadAX promoters
  • Cyclic AMP-CRP binding sites
  • Positive control
  • Escherichia coli