Nitrous oxide reductase copper

Nitric oxide reductase (P) Nitrous oxide reductase (P) Ascorbate oxidase (P) Cytochrome oxidase (PM) Copper ATPase pumps (PM)... 

Figure 3.13 Structure of the Cuz centre in nitrous oxide reductase. The central sulfide interacts with all four copper atoms. (From Rees, 2002. Copyright 2002 Annual Reviews.)...

Nitrite reductases and nitrous oxide reductases are relatively newly found copper-containing proteins involved in bacterial denitrification. N2O reductase may bear a relationship to cytochrome oxidase and, indeed, parallels it somewhat in function, being the terminal electron acceptor in its pathway. 

A new representative of a multicopper cluster in a protein is Cuz in nitrous oxide reductase. As was discussed above this enzyme contains a binuclear CuA centre as in COX. While the latter in addition has CuB in the form of a copper-heme group, N20 reductase has Cuz which is the site of dinitrogen formation from the substrate N20. Recently a central inorganic sulfide has been found as a ligand to copper and multiple forms of Cuz were detected in the enzyme from Paracoccus pantotrophus.134 More recently a tetranuclear copper cluster with X-S bridges was proposed as structure for Cuz..135... 

Reduction of N20 to N2 by bacteria (Eq. 18-30, step d) is catalyzed by the copper-containing nitrous oxide reductase. The purple enzyme is a dimer of 66-kDa subunits, each containing four atoms of Cu.353 It has spectroscopic properties similar to those of cytochome c oxidase and a dinuclear copper-thiolate center similar to that of CuA in cytochrome c oxidase (p. 1030). 

Figure 5.1 Schematic representations of selected active sites of the copper proteins plastocyanin [56] (type 1, a) galactose oxidase [57] (type 2, b) oxy hemocyanin [58] (type 3, c) ascorbate oxidase [10] (type 4, or multicopper site, d) nitrous oxide reductase [59] (CuA site, e) cytochrome c oxidase [15]...

This type of active site is also known as a mixed-valence copper site. Similarly to the type 3 site, it contains a dinuclear copper core, but both copper ions have a formal oxidation state of +1.5 in the oxidized form. This site exhibits a characteristic seven-line pattern in the EPR spectra and is purple colored. Both copper ions have a tetrahedral geometry and are bridged by two sulfur atoms of two cysteinyl residues. Each copper ion is also coordinated by a nitrogen atom from a histidine residue. The function of this site is long-range electron transfer, and it can be found, for example, in cytochrome c oxidase [12-14], and nitrous oxide reductase (Figure 5.1 e). 

The Cuz active site consists of four copper ions, arranged in a distorted tetrahedron and coordinated by seven histidine residues and one hydroxide anion. This site was detected in nitrous oxide reductase [16, 17] (Figure 5.1g) and is involved in the reduction of N20 to N2. The copper ions in the tetranuclear cluster are bridged by an inorganic sulfur ion [18], which until recently was believed to be a hydroxide anion. Three copper ions are coordinated by two histidine residues, whereas the fourth is coordinated by only one, thus leaving a binding site for the substrate. 

Fig. 11. The binuclear CuA (A) and tetranuclear CuZ (B) copper-binding sites of nitrous oxide reductase from Pseduomonas nautica (PDB Accession Code IQNl). The sulfur atom in the tetranuclear copper site is marked with an S.

Figure 2 The dissimilatory denitrification pathway. NO3 is reduced to NO2 by a membrane-bound or periplasmic nitrate reductase(NaR). N02 is reduced to NO by either a cytochrome cdi or copper nitrite reductase (NiR). NO is reduced to N2O by nitric oxide reductase (NOR). N2O is reduced to N2 by nitrous oxide reductase (N2OR). Electron transport from uhiquinol (UQH2) at NaR and the cyt hcj complex is coupled to generation of a proton gradient...

The substrate-binding site in nitrous oxide reductase is a dinuclear copper unit that, in some states, also exists as a mixed-valence species (160y, this center does not show a seven-line EPR spectrum and is similar to the (Class II) mixed-valence derivatives of hemocyanin (156). 

A comparative study of the metal centers in cytochrome c oxidase from several bacterial sources, including Thermus thermophilus and P. denitrificans, using EPR and MCD spectroscopy has established that in both cases cytochrome a is liganded by two histidine oxidases and the Cua center is identical to that in bovine cytochrome c oxidase (105, 106). The properties of the cytochrome Os/Cub dimer have not been established to be identical, although ferrocytochrome 03 is high-spin ferrous, as expected. Recent studies of the MCD properties of the Cua center in cytochrome c oxidase and a copper center in nitrous oxide reductase (107,108) show that the two centers are virtually identical. The evidence from the EPR hyperfine structure of the copper center in nitrous oxide reductase suggests that the center in this enzyme is a mixed-valence Cu(I)/Cu(II) dimer, which raises the interesting prospect that the Cua center in cytochrome c oxidase is also a dimeric copper species. 

Fujiwara and Fukumori, 1996). Nitric oxide reductase is also known, which lacks heme C and uses quinol as the electron donor (Suharti et al., 2001 de Vries et al., 2003). The cytochrome ebb-type enzyme has a molecular structure similar to the structure of the Cub binding portion in cytochrome c oxidase (Saraste and Castresana, 1994 Van der Oost et al., 1994 Zumft et al., 1994). Moreover, quinol NO reductase from Bacillus azotoformans is known to contain Cua. Nitrous oxide is further reduced to nitrogen gas (N2) by the catalysis of nitrous oxide reductase (N20 reductase) which is a multi-copper protein (Zumft and Matsubara, 1982). The structure of the copper-binding portion in the enzyme has been reported also to be similar to the structure of the Cua binding portion of cytochrome c oxidase (Chamock et al., 2000). 

Figure 42 Ribbon diagram of a monomer of nitrous oxide reductase from Achromobacter cycloclastes showing two domains, a C-terminai cupredoxin domain (ieft, in iighter shade) carrying the Cua center (dark biue spheres) and the N-terminai /3-propeiier domain (right, darker shade) with the cataiytic Cuz center in which copper and inorganic S ions are shown as iight biue and dark yeiiow spheres. Reproduced from R. R. Eady S. V. Antonyuk S. S. Hasnain, in Handbook of Metalloproteins, A. Messerschmidt, Ed. John Wiiey Sons Chichester, 2007 Voi. 4, pp 1-15 with permission from John Wiiey Sons.

Anaerobic Growth. In the absence of molecular oxygen, electron flow is dependent on the availability of N-oxides that may serve as alternative electron acceptors 8,10,44, 45, 46). At the shift from oxygen limitation to anaerobiosis, the makeup of the respiratory network drastically changes. The expression of nitrate, nitrite, nitric oxide, and nitrous oxide reductases as well as of the blue copper-containing electron carrier pseudoazurin is established, and in a concerted action these terminal oxidoreductases couple the removal of electrons from the respiratory network to the reduction of the corresponding... 

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