Heme axial ligation by the highly conserved His residues in helix II of cytochrome b (NarI) of Escherichia coli nitrate reductase A.
Optical spectroscopy and EPR studies confirm the existence of two b-type hemes in the NarI subunit (cytochrome bnr) of the membrane-bound nitrate reductase (NarGHI) of Escherichia coli. Replacement of His-56 by Arg and His-66 by Tyr results in the loss of the high-potential heme and of the low-potential heme, respectively. These data support the assignment of the axial ligands to the low-potential heme (His-66 and His-187) and to the high-potential heme (His-56 and His-205). This pairing is consistent with the model proposed for NarI of the nitrate reductase of Thiosphaera pantotropha (Berks, B. C., Page, M. D., Richardson, D. J. , Reilly, A., Cavill, A., Outen, F., and Ferguson, S. J. (1995) Mol. Microbiol. 15, 319-331) in which the two bis-histidine ligated hemes are coordinated by conserved His residues of helix II and V. EPR and optical studies suggest that the low-potential heme (Em,7 = +17 mV) and the high-potential heme (Em,7 = +122 mV) are located near the periplasmic side and the cytoplasmic side of the membrane, respectively. Moreover, correct insertion of both hemes into NarI requires anchoring to NarGH.
Magalon
A
A
Laboratoire de Chimie Bactérienne, IBSM, CNRS, 31 chemin Joseph Aiguier 13402 Marseille cedex 8 France.
Lemesle-Meunier
D
D
Rothery
R A
RA
Frixon
C
C
Weiner
J H
JH
Blasco
F
F
D003080:Cold Temperature; D017124:Conserved Sequence; D003573:Cytochrome b Group / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000201:enzymology*; D006418:Heme / Q000378:metabolism*; D006639:Histidine / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000378:metabolism*; D011199:Potentiometry; D017433:Protein Structure, Secondary; D013054:Spectrophotometry, Atomic
D003573:Cytochrome b Group; D006418:Heme; D006639:Histidine; D009565:Nitrate Reductases; D050901:Nitrate Reductase