Molecular genetic analysis of an FNR-dependent anaerobically inducible Escherichia coli promoter.
From the effects of 13 deletions and three linker-scanner mutations at the Escherichia coli nirB promoter we have located sequences necessary for FNR-dependent induction of activity by anaerobiosis and further nitrite-dependent stimulation of expression. We describe a nirB promoter derivative that allows the cloning of 'cassettes' carrying different FNR-binding sequences and experiments in which a number of point mutations were introduced into these sequences. FNR-dependent stimulation of expression from the nirB promoter is critically dependent on the location of the FNR-binding site, and deletion or insertion of one base pair is sufficient to disrupt promoter function. We have transferred a number of cassette FNR-binding sequences from the nirB promoter to the unrelated melR promoter. The insertion of FNR-binding sequences at the melR promoter is sufficient to confer fnr-dependency on expression. However expression from these hybrid promoters is not as efficiently repressed during aerobic growth, suggesting that the function of bound FNR is dependent on the sequence context of the FNR-binding sequence.
Bell
A I
AI
School of Biochemistry, University of Birmingham, UK.
Cole
J A
JA
Busby
S J
SJ
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D007506:Iron-Sulfur Proteins; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism*
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D014157:Transcription Factors