The nac (nitrogen assimilation control) gene from Klebsiella aerogenes.
The Klebsiella aerogenes nac gene, whose product is necessary for nitrogen regulation of a number of operons, was identified and its DNA sequence determined. The nac sequence predicted a protein a 305 amino acids with a strong similarity to members of the LysR family of regulatory proteins, especially OxyR from Escherichia coli. Analysis of proteins expressed in minicells showed that nac is a single-gene operon whose product has an apparent molecular weight of about 32 kDa as measured in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immediately downstream from nac is a two-gene operon, the first gene of which encodes another member of the LysR family. Upstream from nac is a tRNAAsn gene transcribed divergently from nac. About 60 bp upstream from the nac open reading frame lies a sequence nearly identical to the consensus for sigma 54-dependent promoters, with the conserved GG and GC nucleotides at -26 and -14 relative to the start of transcription. About 130 bp farther upstream (at -153 relative to the start of transcription) is a sequence nearly identical to the transcriptional activator NTRC-responsive enhancer consensus. Another weaker NTRC-binding site is located adjacent to this site (at -133 relative to the start of transcription). Thus, we propose that nac is transcribed by RNA polymerase carrying sigma 54 in response to the nitrogen regulatory (NTR) system. A transposon located between the promoter and the nac ORF prevented NTR-mediated expression of nac, supporting this identification of the promoter sequence. The insertion of over 5 kb of transposon DNA between the enhancer and its target promoter had only a weak effect on enhancer-mediated regulation, suggesting that enhancers may be able to act at a considerable distance on the bacterial chromosome.
Schwacha
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Department of Biology, University of Michigan, Ann Arbor 48109-1048.
Bender
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002874:Chromosome Mapping; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics*; D005816:Genetic Complementation Test; D007711:Klebsiella pneumoniae / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009584:Nitrogen / Q000172:deficiency / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D012097:Repressor Proteins; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000235:genetics*
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C072712:NAC protein, bacteria; C489526:Nac protein, E coli; D012097:Repressor Proteins; D014157:Transcription Factors; C073470:LysR protein, Bacteria; D009584:Nitrogen