Transcription activation by the Escherichia coli cyclic AMP receptor protein. Receptors bound in tandem at promoters can interact synergistically

J Mol Biol. 1994 Aug 19;241(3):341-52. doi: 10.1006/jmbi.1994.1511.

Abstract

Starting with a semi-synthetic Escherichia coli promoter with a binding site for the cyclic AMP receptor protein (CRP) centred between base-pairs 41 and 42 upstream from the transcription start site, a second upstream CRP-binding site, centred between base-pairs 90 and 91, was introduced. CRP binding to this second upstream site results in a several-fold greater stimulation of CRP-dependent transcription initiation, compared to activation at the starting promoter with just one CRP-binding site. Activation of transcription by the upstream CRP molecule is blocked by the HL159 substitution, suggesting that the upstream-bound CRP makes a direct contact with RNA polymerase. Footprinting experiments suggest that RNA polymerase contacts the promoter DNA between the two CRP-binding sites, most likely due to interactions involving the C-terminal part of the alpha subunit. Synergy between tandem bound CRP molecules in transcription activation requires that the two CRP-binding sites be separated by around 40 or 50 base-pairs, but is not found at intermediate spacings. An experiment in which the upstream CRP-binding site is replaced by a site for the related transcription factor, FNR, shows that heterologous synergistic interactions between FNR and CRP are possible.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / metabolism
  • Base Sequence
  • Binding Sites
  • Cyclic AMP Receptor Protein / metabolism*
  • Cyclic AMP Receptor Protein / physiology
  • DNA, Bacterial / metabolism*
  • DNA-Directed RNA Polymerases / metabolism
  • Escherichia coli / genetics*
  • Escherichia coli Proteins*
  • Iron-Sulfur Proteins*
  • Models, Biological
  • Molecular Sequence Data
  • Promoter Regions, Genetic / genetics*
  • RNA, Bacterial / biosynthesis
  • RNA, Messenger / biosynthesis
  • Transcription Factors / metabolism
  • Transcription, Genetic
  • Transcriptional Activation / physiology*

Substances

  • Bacterial Proteins
  • Cyclic AMP Receptor Protein
  • DNA, Bacterial
  • Escherichia coli Proteins
  • FNR protein, E coli
  • Iron-Sulfur Proteins
  • RNA, Bacterial
  • RNA, Messenger
  • Transcription Factors
  • DNA-Directed RNA Polymerases