Influence of the location of the cAMP receptor protein binding site on the geometry of a transcriptional activation complex in Escherichia coli

Biochemistry
P EichenbergerJ Geiselmann

Abstract

The interactions between the cAMP receptor protein (CRP) and RNA polymerase during transcriptional activation at the Escherichia coli malT promoter have been analyzed using a combination of footprinting methods. We show that a closed complex is formed at this promoter in the absence of activator and that CRP merely stabilizes the open complex. The alpha-subunits of the RNA polymerase are involved in this effect as shown by KMnO4 footprinting. The open complex formed in the presence of CRP is structurally identical to the one found at a CRP-independent promoter up-mutant. UV-laser footprinting yields distinct signals for the different protein-DNA interactions within the complex and for interactions between CRP and RNA polymerase. We monitor these signals in promoter variants that place the CRP binding site at different distances upstream of the start site of transcription. Signals within the core promoter region, as well as those located just upstream of the -35 hexamer, are unaffected by the position of the CRP binding site. Contacts of RNA polymerase with the upstream promoter region change in a mutant RNA polymerase containing a truncated alpha-subunit. We conclude that at least one of the alpha-subunits of RNA polymerase bin...Continue Reading

References

May 20, 1991·Journal of Molecular Biology·S S Zinkel, D M Crothers
Dec 1, 1983·Journal of Bacteriology·C Chapon, A Kolb
Jan 14, 1994·Journal of Molecular Biology·A KumarA Ishihama
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Citations

Mar 6, 1999·Current Opinion in Microbiology·V A Rhodius, S J Busby
Aug 19, 1997·Proceedings of the National Academy of Sciences of the United States of America·P EichenbergerJ Geiselmann
Oct 29, 2014·Nature Communications·W Ryan WillFerric C Fang
Nov 5, 1999·Journal of Molecular Biology·S Busby, R H Ebright

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