Functional roles for the GerE-family carboxyl-terminal domains of nitrate response regulators NarL and NarP of Escherichia coli K-12.
NarL and NarP are paralogous response regulators that control anaerobic gene expression in response to the favoured electron acceptors nitrate and nitrite. Their DNA-binding carboxyl termini are in the widespread GerE-LuxR-FixJ subfamily of tetrahelical helix-turn-helix domains. Previous biochemical and crystallographic studies with NarL suggest that dimerization and DNA binding by the carboxyl-terminal domain (CTD) is inhibited by the unphosphorylated amino-terminal receiver domain. We report here that NarL-CTD and NarP-CTD, liberated from their receiver domains, activated transcription in vivo from the class II napF and yeaR operon control regions, but failed to activate from the class I narG and fdnG operon control regions. Alanine substitutions were made to examine requirements for residues in the NarL DNA recognition helix. Substitutions for Val-189 and Arg-192 blocked DNA binding as assayed both in vivo and in vitro, whereas substitution for Arg-188 had a strong effect only in vivo. Similar results were obtained with the corresponding residues in NarP. Finally, Ala substitutions identified residues within the NarL CTD as important for transcription activation. Overall, results are congruent with those obtained for other GerE-family members, including GerE, TraR, LuxR and FixJ.
Lin
Alice V
AV
Biochemistry and Molecular Biology Graduate Group, University of California, Davis, CA 95616-8665, USA.
Stewart
Valley
V
Department of Microbiology, University of California, Davis, CA 95616-8665, USA.|Biochemistry and Molecular Biology Graduate Group, University of California, Davis, CA 95616-8665, USA.
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009566:Nitrates / Q000378:metabolism; D009876:Operon; D011485:Protein Binding; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D009566:Nitrates; C081466:narP protein, E coli; C069780:NarL protein, E coli