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Related Experiment Videos

Specific binding of PapI to Lrp-pap DNA complexes

L S Kaltenbach1, B A Braaten, D A Low

  • 1Department of Pathology, University of Utah Health Sciences Center, Salt Lake City 84132, USA.

Journal of Bacteriology
|November 1, 1995
PubMed
Summary

Pyelonephritis-associated pili (Pap) phase variation relies on DNA methylation and regulatory proteins. PapI protein facilitates the switch between transcriptionally active (ON) and inactive (OFF) states by interacting with leucine-responsive regulatory protein (Lrp).

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • Pyelonephritis-associated pili (Pap) expression exhibits phase variation, switching between transcriptionally active (ON) and inactive (OFF) states.
  • This phase variation is regulated by the leucine-responsive regulatory protein (Lrp) binding to specific DNA regions (GATC-I and GATC-II) within the pap operon.
  • Deoxyadenosine methylase sites within these GATC regions are crucial, as their methylation status influences Lrp binding and thus Pap expression.

Purpose of the Study:

  • To elucidate the role of the PapI protein in regulating Pap phase variation.
  • To investigate the interaction between PapI and Lrp in controlling transcription states.
  • To understand how PapI influences Lrp binding dynamics at the papBA promoter.

Main Methods:

Related Experiment Videos

  • Gel shift analysis using radiolabeled PapI to assess binding to Lrp-pap DNA complexes.
  • Protein cross-linking studies to determine direct interactions between Lrp and PapI.
  • Investigating the effect of PapI on Lrp binding localization at pap regulatory regions.

Main Results:

  • PapI binds specifically to Lrp-pap DNA complexes but shows weak binding to free Lrp.
  • Lrp binding shifts from overlapping the papBA promoter (OFF state) to upstream regions (ON state) upon PapI addition.
  • Protein cross-linking confirms a direct physical interaction between Lrp and PapI.

Conclusions:

  • PapI plays a critical role in mediating the switch between OFF and ON transcriptional states of Pap.
  • PapI likely functions by binding to Lrp, thereby altering Lrp's affinity for the pap GATC-I and GATC-II regulatory regions.
  • This interaction provides a mechanism for precise control over Pap phase variation, essential for uropathogenic E. coli virulence.