The regulator of fumarate and nitrate reduction (FNR) protein of is an oxygen-responsive transcription regulator that acts mainly to activate the transcription of genes associated with anaerobic energy generation during periods of oxygen starvation. The gene of the swine pathogen encodes an FNR homologue, HlyX, which can complement the anaerobic respiratory deficiencies of an mutant. However, FNR and HlyX have distinct but overlapping regulons because during anaerobic incubation, -expressing K-12 strains produce an otherwise latent haemolysin. The gene encoding the ‘latent’ haemolysin has been designated and analysis of the promoter region by DNase I footprinting reveals the presence of an FNR- (HlyX-) binding site. Anaerobic expression of an reporter was 6.5-fold higher in compared to -expressing cells. Both FNR and HlyX recruited RNA polymerase to the promoter but formed different ternary complexes. One major transcript (tsp1) initiating at 78.5 bp downstream of the FMR-binding site and four minor transcripts initiating at 73.5 (tsp2), 71.5 (tsp3), 63.5 (tsp4) and 62.5 (tsp5) bp from the FNR site were detected. From the position of the FNR box relative to the transcript starts, is expressed from a Class I FNR-regulated promoter. Substitution of selected FNR amino acids with the residues found in the equivalent positions in HlyX indicated that Activating Region 1 (AR1) of FNR forms a surface encompassing β to β and that the AR1 contact at Class I promoters is different to that at Class II promoters, although the same surface is involved. The FNR variant, FNR-A225T, combined the properties of FNR (good activation from Class II promoters) and HlyX (good activation of Class I promoters) and conferred the haemolytic phenotype.

Keyword(s): Class I promoter , FNR , haemolysin , HlyE and HlyX

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