Reorganisation of an RNA polymerase-promoter DNA complex for DNA melting

Patricia C. Burrows, Konstantin Severinov, Martin Buck, Siva R. Wigneshweraraj

Research output: Contribution to journalArticlepeer-review

33 Scopus citations

Abstract

Sigma factors, the key regulatory components of the bacterial RNA polymerase (RNAP), direct promoter DNA binding and DNA melting. The σ54-RNAP forms promoter complexes in which DNA melting is only triggered by an activator and ATP hydrolysis-driven reorganisation of an initial σ54-RNAP-promoter complex. We report that an initial bacterial RNAP-DNA complex can be reorganised by an activator to form an intermediate transcription initiation complex where full DNA melting has not yet occurred. Using σ54 as a chemical nuclease we now show that the reorganisation of the initial σ54-RNAP-promoter complex occurs upon interaction with the activator at the transition point of ATP hydrolysis. We demonstrate that this reorganisation event is an early step in the transcription initiation pathway that occurs independently of RNAP parts normally associated with stable DNA melting and open complex formation. Using photoreactive DNA probes, we provide evidence that within this reorganised σ54-RNAP-promoter complex, DNA contacts across the 'to be melted' sequences are made by the σ54 subunit. Strikingly, the activator protein, but not core RNAP subunits, is close to these DNA sequences.

Original languageEnglish (US)
Pages (from-to)4253-4263
Number of pages11
JournalEMBO Journal
Volume23
Issue number21
DOIs
StatePublished - Oct 27 2004

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)
  • Molecular Biology
  • Neuroscience(all)

Keywords

  • AAA activator
  • DNA cleavage
  • Photo-crosslinking
  • RNA polymerase
  • Sigma factors

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