title: Analysis of the nitric oxide-sensing non-heme iron center in the NorR regulatory protein.
abstract: The NorR regulatory protein senses nitric oxide (NO) to activate genes required for NO detoxification under anaerobic and microaerobic conditions in Escherichia coli. NorR belongs to the sigma(54)-dependent family of transcriptional activators and contains an N-terminal regulatory GAF (cGMP phosphodiesterase, adenylate cyclase, FhlA) domain that controls the ATPase activity of the central AAA+ domain to regulate productive interactions with sigma(54). Binding of NO to a non-heme iron center in the GAF domain results in the formation of a mononitrosyl-iron complex and releases intramolecular repression of the AAA+ domain to enable activation of transcription. In this study, we have further characterized NorR spectroscopically and substituted conserved residues in the GAF domain. This analysis, in combination with structural modeling of the GAF domain, has identified five candidate ligands to the non-heme iron and suggests a model in which the metal ion is coordinated in a pseudo-octahedral environment by three aspartate residues, an arginine, and a cysteine.
authors.affiliation: Department of Molecular Microbiology, John Innes Centre, Colney, Norwich Research Park, Norwich NR4 7UH, United Kingdom.
authors.forename: Nicholas P
authors.initials: NP
authors.lastname: Tucker
authors.forename: Benoît
authors.initials: B
authors.lastname: D'Autréaux
authors.forename: Faridoon K
authors.initials: FK
authors.lastname: Yousafzai
authors.forename: Shirley A
authors.initials: SA
authors.lastname: Fairhurst
authors.forename: Stephen
authors.initials: S
authors.lastname: Spiro
authors.forename: Ray
authors.initials: R
authors.lastname: Dixon
mesh_terms: D000595:Amino Acid Sequence; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008658:Inactivation, Metabolic; D007501:Iron / Q000032:analysis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000378:metabolism* / Q000633:toxicity; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013053:Spectrophotometry; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D029968:Escherichia coli Proteins; C504602:NorR protein, E coli; D011994:Recombinant Proteins; D015534:Trans-Activators; D009569:Nitric Oxide; D007501:Iron