Direct inhibition by nitric oxide of the transcriptional ferric uptake regulation protein via nitrosylation of the iron.
Ferric uptake regulation protein (Fur) is a bacterial global regulator that uses iron as a cofactor to bind to specific DNA sequences. The function of Fur is not limited to iron homeostasis. A wide variety of genes involved in various mechanisms such as oxidative and acid stresses are under Fur control. Flavohemoglobin (Hmp) is an NO-detoxifying enzyme induced by NO and nitrosothiol compounds. Fur recently was found to regulate hmp in Salmonella typhimurium, and in Escherichia coli, the iron-chelating agent 2,2'-dipyridyl induces hmp expression. We now establish direct inhibition of E. coli Fur activity by NO. By using chromosomal Fur-regulated lacZ reporter fusion in E. coli, Fur activity is switched off by NO at micromolar concentration. In vitro Fur DNA-binding activity, as measured by protection of restriction site in aerobactin promoter, is directly sensitive to NO. NO reacts with Fe(II) in purified FeFur protein to form a S = 12 low-spin FeFur-NO complex with a g = 2.03 EPR signal. Appearance of the same EPR signal in NO-treated cells links nitrosylation of the iron with Fur inhibition. The nitrosylated Fur protein is still a dimer and is stable in anaerobiosis but slowly decays in air. This inhibition probably arises from a conformational switch, leading to an inactive dimeric protein. These data establish a link between control of iron metabolism and the response to NO effects.
D'Autreaux
Benoit
B
Laboratoire de Physicochimie des Métaux en Biologie (Formation de Recherche en Evolution-Université Joseph Fourier-Commissariat à l'Energie Atomique-Centre National de la Recherche Scientifique no. 2427), Grenoble, F-38054 Grenoble Cedex 9, France.
Touati
Daniele
D
Bersch
Beate
B
Latour
Jean-Marc
JM
Michaud-Soret
Isabelle
I
D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000502:physiology; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli; D007501:Iron / Q000378:metabolism*; D009569:Nitric Oxide / Q000502:physiology*; D011487:Protein Conformation; D012097:Repressor Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000502:physiology; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D012097:Repressor Proteins; C068477:ferric uptake regulating proteins, bacterial; D009569:Nitric Oxide; D007501:Iron