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The Klebsiella pneumoniae nifJ promoter: analysis of promoter elements regulating activation by the Nif A promoter
Author(s) -
Charlton Wayne,
Can Wendy,
Buck Martin
Publication year - 1993
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.1993.tb01192.x
Subject(s) - klebsiella pneumoniae , biology , promoter , microbiology and biotechnology , computational biology , gene , genetics , escherichia coli , gene expression
Summary The nifJ and nifH promoters of Klebsiella pneumoniae are divergently transcribed σ; 54 ‐dependent promoters that are positively activated by the NifA protein. NifA binds to upstream activator sequences (UASs), usually located 60‐200 bp upstream of the start of transcription. Bound NifA is presented to the RNA polymerase‐σ 54 complex (Eσ 54 ) via DNA loop formation, mediated by the binding of integration host factor protein (IHF) between Eσ 54 and NifA. The nifJ promoter sequence contains three potential NifA binding sites (UAS1, 2 and 3) and two potential RNA polymerase‐σ 54 ‐binding sites (downstream promoter elements, DPEs 1 and 2). DPE2 is located 420 bp into the coding region and DPE1 overlaps UAS1 by 5 bp. Mutational and footprinting analyses have shown efficient activation of the nifJ promoter requires that NifA is bound at UAS 2 and 3. Transcription is initiated at DPE1. Only a weak interaction of NifA with the UAS overlapping DPE1 was detected. Footprints demonstrated that Eσ 54 forms a closed complex at DPE1 but not DPE2 and that bound Eσ 54 closely approaches the −15 region of DPE1. Stimulation of nifJ promoter activity by IHF was not as great as that observed for other nif promoters. In the absence of IHF nifH promoter sequences stimulated activation of the nifJ promoter. This appeared to require NifA bound at the nifH UAS. Thus, one additional role of IHF may be to partition NifA between the two promoters by constraining the topology of the DNA.