Mapping σ54-RNA polymerase interactions at the -24 consensus promoter element

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

Research output: Contribution to journalArticlepeer-review

29 Citations (Scopus)

Abstract

The σ54 promoter specificity factor is distinct from σ70-type factors. The σ54-RNA polymerase binds to promoters with conserved sequence elements at -24 and -12 and utilizes specialized enhancer-binding activators to convert, through an ATP-dependent process, closed promoter complexes to open promoter complexes. The interface between σ54-RNA polymerase and promoter DNA is poorly characterized, contrasting with σ70. Here, σ54 was modified with strategically positioned cleavage reagents to provide physical evidence that the highly conserved RpoN box motif of σ54 is close to and may therefore interact with the consensus -24 promoter element. We show that the spatial relationship between the σ54-RNA polymerase and the -24 promoter element remains unchanged during closed to open complex conversion and transcription initiation but changes during the early elongation phase. In contrast, the spatial relationship between σ54-RNA polymerase and the consensus -12 promoter element changes upon conversion of the closed promoter complex to an open one. We provide evidence that some -12 promoter region-σ54 interactions are dependent upon either the core RNA polymerase or a fork junction DNA structure at the -12-position, indicating that DNA fork junctions can substitute for core RNAP. We also show the β-subunit flap domain contributes to different sets of σ-promoter DNA interactions at σ54- and σ70-dependent promoters.

Original languageEnglish
Pages (from-to)29728-29743
Number of pages16
JournalJournal of Biological Chemistry
Volume278
Issue number32
DOIs
Publication statusPublished - 8 Aug 2003
Externally publishedYes

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