title: Characterisation of [Cu4S], the catalytic site in nitrous oxide reductase, by EPR spectroscopy.
abstract: The enzyme nitrous oxide reductase (N(2)OR) has a unique tetranuclear copper centre [Cu(4)S], called Cu(Z), at the catalytic site for the two-electron reduction of N(2)O to N(2). The X- and Q-band EPR spectra have been recorded from two forms of the catalytic site of the enzyme N(2)OR from Paracoccus pantotrophus, namely, a form prepared anaerobically, Cu(Z), that undergoes a one-electron redox cycle and Cu(Z)*, prepared aerobically, which cannot be redox cycled. The spectra of both species are axial with that of Cu(Z) showing a rich hyperfine splitting in the g||-region at X-band. DFT calculations were performed to gain insight into the electronic configuration and ground-state properties of Cu(Z) and to calculate EPR parameters. The results for the oxidation state [Cu(+1)(3)Cu(+2)(1)S](3+) are in good agreement with values obtained from the fitting of experimental spectra, confirming the absolute oxidation state of Cu(Z). The unpaired spin density in this configuration is delocalised over four copper ions, thus, Cu(I) 20.1%, Cu(II) 9.5%, Cu(III) 4.8% and Cu(IV) 9.2%, the mu(4)-sulfide ion and oxygen ligand. The three copper ions carrying the highest spin density plus the sulfide ion lie approximately in the same plane while the fourth copper ion is perpendicular to this plane and carries only 4.8% spin density. It is suggested that the atoms in this plane represent the catalytic core of Cu(Z), allowing electron redistribution within the plane during interaction with the substrate, N(2)O.
authors.affiliation: Centre for Metalloprotein Spectroscopy and Biology, Schools of Chemical Sciences and Pharmacy, University of East Anglia, Norwich NR4 7TJ, UK.
authors.forename: Vasily S
authors.initials: VS
authors.lastname: Oganesyan
authors.forename: Tim
authors.initials: T
authors.lastname: Rasmussen
authors.forename: Shirley
authors.initials: S
authors.lastname: Fairhurst
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Thomson
mesh_terms: D001665:Binding Sites; D020134:Catalytic Domain; D003300:Copper / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D004583:Electrons; D008958:Models, Molecular; D008968:Molecular Conformation; D015394:Molecular Structure; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D042084:Paracoccus pantotrophus / Q000201:enzymology; D013440:Sulfides / Q000737:chemistry*
chemical_list: D013440:Sulfides; D003300:Copper; D010088:Oxidoreductases; C022763:nitrous oxide reductase