Location and orientation of an activating region in the Escherichia coli transcription factor, FNR

We have characterized a number of mutations in fnr that interfere with FNR-dependent transcription activation at two promoters where the FNR-binding site is centred around 41 1/2 bp upstream from the transcription start site. The substituted residues in all but one of these FNR mutants are clustered around a presumed surface-exposed beta-turn containing G85 which, we suggest, forms an activating region that contacts RNA polymerase at these promoters. Using the 'oriented heterodimers' method described elsewhere, we show that this activating region on the promoter-proximal subunit of the FNR dimer is sufficient to activate transcription initiation. In contrast, this region is not essential for activation of a third FNR-dependent promoter where the FNR-binding site is centred at 61 1/2 bp upstream from the transcription start site. However, a substitution at S73 interferes with FNR-dependent activation at both this promoter and promoters in which the FNR site is located at 41 1/2 bp from the transcript start, suggesting that FNR may contain a second activating region.