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Positioning of σ S , the stationary phase σ factor, in Escherichia coli RNA polymerase–promoter open complexes
Author(s) -
Colland Frédéric,
Fujita Nobuyuki,
Kotlarz Denise,
Bown Jonathan A.,
Meares Claude F.,
Ishihama Akira,
Kolb Annie
Publication year - 1999
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/18.14.4049
Subject(s) - biology , escherichia coli , sigma factor , stationary phase , rna polymerase , microbiology and biotechnology , polymerase , genetics , gene , chemistry , chromatography
The σ S subunit of RNA polymerase is the master regulator of the general stress response in Escherichia coli and is required for promoter recognition of many stationary phase genes. We have analysed open complexes of Eσ S RNA polymerase, using σ S derivatives carrying single cysteine residues at nine different positions to which the reagent FeBABE has been tethered. All holoenzymes but one formed transcriptionally active open complexes at three different promoters ( osmY , gal P1 and lac UV5). The chemical nuclease FeBABE can cleave DNA in proximity to the chelate. The overall cutting pattern of Eσ S open complexes does not depend on the nature of the promoter and is similar to that obtained with Eσ 70 , but extends towards the downstream part of the promoter. The strongest cleavages are observed with FeBABE positioned on cysteines in regions 2.2 to 3.1. In contrast to σ 70 , region 2.1 of σ S appears to be far from DNA. Region 4.2 of σ S appears less accessible than its counterpart in σ 70 and FeBABE positioned in the turn of the helix–turn–helix (HTH) motif in region 4.2 reacts only weakly with the −35 promoter element. This provides a structural basis for the minor role of the −35 sequence in σ S ‐dependent promoter recognition.