Activation of transcription initiation from the nac promoter of Klebsiella aerogenes.
The nac gene of Klebsiella aerogenes encodes a bifunctional transcription factor that activates or represses the expression of several operons under conditions of nitrogen limitation. In experiments with purified components, transcription from the nac promoter was initiated by sigma 54 RNA polymerase and was activated by the phosphorylated form of nitrogen regulator I (NRI) (NtrC). The activation of the nac promoter required a higher concentration of NRI approximately P than did the activation of the Escherichia coli glnAp2 promoter, and both the promoter and upstream enhancer element contributed to this difference. The nac promoter had a lower affinity for sigma 54 RNA polymerase than did glnAp2, and uninitiated competitor-resistant transcription complexes formed at the nac promoter decayed to competitor-sensitive complexes at a greater rate than did similar complexes formed at the glnAp2 promoter. The nac enhancer, consisting of a single high-affinity NRI-binding site and an adjacent site with low affinity for NRI, was less efficient in stimulating transcription than was the glnA enhancer, which consists of two adjacent high-affinity NRI-binding sites. When these binding sites were exchanged, transcription from the nac promoter was increased and transcription from the glnAp2 promoter was decreased at low concentrations of NRI approximately P. Another indication of the difference in the efficiency of these enhancers is that although activation of a nac promoter construct containing the glnA enhancer was relatively insensitive to subtle alterations in the position of these sites relative to the position of the promoter, activation of the natural nac promoter or a nac promoter construct containing only a single high-affinity NRI approximately P binding site was strongly affected by subtle alterations in the position of the NRI approximately P binding site(s), indicating a face-of-the-helix dependency for activation.
Feng
J
J
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor 48109, USA.
Goss
T J
TJ
Bender
R A
RA
Ninfa
A J
AJ
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001667:Binding, Competitive; D004269:DNA, Bacterial / Q000378:metabolism; D004278:DNA, Superhelical / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D029968:Escherichia coli Proteins; D005974:Glutamate-Ammonia Ligase / Q000235:genetics; D007711:Klebsiella pneumoniae / Q000235:genetics*; D008969:Molecular Sequence Data; D051922:PII Nitrogen Regulatory Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051877:RNA Polymerase Sigma 54; D012808:Sigma Factor / Q000378:metabolism; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics; D015533:Transcriptional Activation / Q000235:genetics*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C072712:NAC protein, bacteria; C489526:Nac protein, E coli; D051922:PII Nitrogen Regulatory Proteins; D012808:Sigma Factor; D015534:Trans-Activators; D014157:Transcription Factors; C497876:glnG protein, E coli; C497751:rpoN protein, E coli; C404143:PIID regulatory protein, Bacteria; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; C081173:glutamine synthetase I; D005974:Glutamate-Ammonia Ligase