Functional regions of the N-terminal domain of the antiterminator RfaH

Mol Microbiol. 2010 Apr;76(2):286-301. doi: 10.1111/j.1365-2958.2010.07056.x. Epub 2010 Feb 1.

Abstract

RfaH is a bacterial elongation factor that increases expression of distal genes in several long, horizontally acquired operons. RfaH is recruited to the transcription complex during RNA chain elongation through specific interactions with a DNA element called ops. Following recruitment, RfaH remains bound to RNA polymerase (RNAP) and acts as an antiterminator by reducing RNAP pausing and termination at some factor-independent and Rho-dependent signals. RfaH consists of two domains connected by a flexible linker. The N-terminal RfaH domain (RfaH(N)) recognizes the ops element, binds to the RNAP and reduces pausing and termination in vitro. Functional analysis of single substitutions in this domain reported here suggests that three separate RfaH(N) regions mediate these functions. We propose that a polar patch on one side of RfaH(N) interacts with the non-template DNA strand during recruitment, whereas a hydrophobic surface on the opposite side of RfaH(N) remains bound to the beta' subunit clamp helices domain throughout transcription of the entire operon. The third region is apparently dispensable for RfaH binding to the transcription complex but is required for the antitermination modification of RNAP.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Substitution / genetics
  • DNA Mutational Analysis
  • DNA, Bacterial / metabolism
  • DNA-Directed RNA Polymerases / metabolism
  • Escherichia coli / physiology*
  • Escherichia coli Proteins / genetics*
  • Escherichia coli Proteins / metabolism*
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Peptide Elongation Factors / genetics*
  • Peptide Elongation Factors / metabolism*
  • Protein Binding
  • Protein Structure, Quaternary
  • Protein Structure, Tertiary
  • Regulatory Elements, Transcriptional
  • Trans-Activators / genetics*
  • Trans-Activators / metabolism*
  • Transcription, Genetic*

Substances

  • DNA, Bacterial
  • Escherichia coli Proteins
  • Peptide Elongation Factors
  • RfaH protein, E coli
  • Trans-Activators
  • DNA-Directed RNA Polymerases