Copper enzymes nitrite reductase

There are a number of excellent sources of information on copper proteins notable among them is the three-volume series Copper Proteins and Copper Enzymes (Lontie, 1984). A review of the state of structural knowledge in 1985 (Adman, 1985) included only the small blue copper proteins. A brief review of extended X-ray absorption fine structure (EXAFS) work on some of these proteins appeared in 1987 (Hasnain and Garner, 1987). A number of new structures have been solved by X-ray diffraction, and the structures of azurin and plastocyanin have been extended to higher resolution. The new structures include two additional type I proteins (pseudoazurin and cucumber basic blue protein), the type III copper protein hemocyanin, and the multi-copper blue oxidase ascorbate oxidase. Results are now available on a copper-containing nitrite reductase and galactose oxidase. 

As mentioned earlier, the copper containing nitrite reductase is a trimer of identical subunits. In each subunit there is a type 1 copper which acts analogously to the c-type heme in cytochrome cd and thus is the point of entry of electron into the enzyme. The three eatalytic sites have type 2... 

Intramolecular ET between distinct copper centers is part of the catalytic cycles of many copper-containing redox enzymes, such as the multicopper oxidases, ascorbate oxidase, and ceruloplasmin, as well as the copper-containing nitrite reductases. Examination of internal LRET in these proteins is of considerable interest as it may also provide insights into the evolution of selected ET pathways in particular, whether and how the enzymes have evolved in order to optimize catalytic functions. With the increase in the number of known high-resolution 3D structures of transition metal containing redox enzymes, studies of structure-reactivity relationships have become feasible and indeed many have been carried out during the last two decades. 

It is of interest to compare rate constants and activation parameters of the intramolecular ET in Ps-NiR with those determined for analogous intramolecular ET processes in other multicentered redox enzymes. Reversible intramolecular ET reaction between type 1 and 2 sites in the copper-containing nitrite reductases (CuNiR isolated from A. xylosoxidans and A. cycloclastes)... 

Besides dioxygen, nitrogen oxides can serve as electron acceptors in reactions catalyzed by copper enzymes. The copper-containing nitrite reductase (NIR) from denitrifying bacteria such as Achromobaaer, Pseudomonas, or Rhodobacter is part of the dissimilatory metabolic pathway of these bacteria. The enzyme catalyzes the one-electron reduction of NO2 to NO and water according to Equations (14) and (15). 

The blue copper center in cupredoxins is also found in multidomain, multicopper enzymes " such as ascorbate oxidase (AO), laccase (Lc), human ceruloplasmin (Cp), " and a subfamily of copper-containing nitrite reductase. Nitrite reductase (NiR) catalyzes reduction of nitrite (N(32 ) to nitric oxide (NO), a step in the biological dinitrification cycle. Two distinct types of NIR are known multiheme NiR (see Chapter 8.29) and multicopper NiR. The multicopper... 

The copper-containing nitrite reductases (Cu NiR s) also contain two distinct types of chromophore. The best characterized is that from Achromobacter cycloclastes, for which a 2.3 A resolution X-ray structure has been reported (12). The enzyme consists of an a3 trimer of 34.5 kDa subunits, each of which contains two copper atoms in distinct sites. One is an... 

Fig. 6.9 The catalysts for denitrification. Nitrate is reduced by a molybdenum enzyme while nitrite and oxides of nitrogen are reduced today mainly by copper enzymes. However, there are alternatives, probably earlier iron enzymes. The electron transfer bct complex is common to that in oxidative phosphorylation and similar to the bf complex of photosynthesis, while cytochrome c2 is to be compared with cytochrome c of oxidative phosphorylation. These four processes are linked in energy capture via proton (H+) gradients see Figure 6.8(a) and (b) and the lower parts of Fig. 6.9 which show separately the active site of the all iron NO-reductase, and the active site of cytochrome oxidase (02 reductase).

Several copper enzymes will be discussed in detail in subsequent sections of this chapter. Information about major classes of copper enzymes, most of which will not be discussed, is collected in Table 5.1 as adapted from Chapter 14 of reference 49. Table 1 of reference 4 describes additional copper proteins such as the blue copper electron transfer proteins stellacyanin, amicyanin, auracyanin, rusticyanin, and so on. Nitrite reductase contains both normal and blue copper enzymes and facilitates the important biological reaction NO) — NO. Solomon s Chemical Reviews article4 contains extensive information on ligand field theory in relation to ground-state electronic properties of copper complexes and the application of... 

The NO/NO+ and NO/NO- self-exchange rates are quite slow (42). Therefore, the kinetics of nitric oxide electron transfer reactions are strongly affected by transition metal complexes, particularly by those that are labile and redox active which can serve to promote these reactions. Although iron is the most important metal target for nitric oxide in mammalian biology, other metal centers might also react with NO. For example, both cobalt (in the form of cobalamin) (43,44) and copper (in the form of different types of copper proteins) (45) have been identified as potential NO targets. In addition, a substantial fraction of the bacterial nitrite reductases (which catalyze reduction of NO2 to NO) are copper enzymes (46). The interactions of NO with such metal centers continue to be rich for further exploration. 

Nitrite reductase (NAD(P)H) [EC 1.6.6.4] catalyzes the reaction of three NAD(P)H with nitrite to yield three NAD(P)+, NH4OH, and water. Cofactors for this enzyme include FAD, non-heme iron, and siroheme. (2) Nitrite reductase (cytochrome) [EC 1.7.2.1] is a copper-depen-dent system that catalyzes the reaction of nitric oxide with two ferricytochrome c and water to produce nitrite and two ferrocytochrome c. (3) Ferredoxin-nitrite reductase [EC 1.7.7.1], a heme- and iron-dependent enzyme, catalyzes the reaction of ammonia with three oxidized ferredoxin to produce nitrite and three reduced ferredoxin. (4) Nitrite reductase [EC 1.7.99.3] is a copper- and FAD-dependent enzyme that catalyzes the reaction of two nitric oxide with an acceptor substrate and two water to produce two nitrite and the reduced acceptor. 

Two types of dissimilatory nitrite reductases catalyze step b of Eq. 18-30. Some bacteria use a copper-containing enzyme, which contains a type 1 (blue) copper bound to a (3 barrel domain of one subunit and a type 2 copper at the catalytic center. The type 1 copper is thought to receive electrons from the small copper-containing carrier pseudoazurin (Chapter... 

This chapter focuses on the chemistry ofbiomimetic copper nitrosyl complexes relevant to the NO-copper interactions in proteins that are central players in dissimilatory nitrogen oxide reduction (denitrification). The current state of knowledge of NO-copper interactions in nitrite reductase, a key denitrifying enzyme, is briefly surveyed the syntheses, structures, and reactivity of copper nitrosyl model complexes prepared to date are presented and the insight these model studies provide into the mechanisms of denitrification and the structures of other copper protein nitrosyl intermediates are discussed. Emphasis is placed on analysis of the geometric features, electronic structures, and biomimetic reactivity with NO or NOf of the only structurally characterized copper nitrosyls, a dicopper(II) complex bridged by NO and a mononuclear tris(pyrazolyl)hydroborate complex having a Cu(I)-NO formulation. 

Type II copper(II) sites are present in mononuclear copper enzymes such as dioxygenases, monoxygenases, nitrite reductases, and nonblue oxidases. The high molecular weight of most of these enzymes and the unfavorable electron relaxation time of copper ion in the oxidized form have up to now precluded the application of NMR spectroscopy. 

There are four different types of nitrite reductases the copper-containing protein Copper Enzymes in Denitrification and cytochrome cd perform a one-electron rednetion of nitrite to nitric oxide, and are involved in denitrification " the siroheme-containing protein and the cytochrome c ititrite reductase (cNiR) both perform the complete, six-electron reduction, of nitrite to ammonia. The cNiR is present in the y, 5 or e-subclasses of proteobacteria, and is encoded by the nrf operon (nitrite reduction with /ormate), which has different gene composition in the different classes of bacteria, having in common only the gene for the catalytic subunit, ntfA. 

Some nitrite reductases contain iron and copper other enzymes active in these reactions contain manganese. Reactions catalyzed by copper and iron enzymes with NO, N2O, and N2 as products have also been reported. 

Direct reaction of NO with enzymes has been shown for cytochrome c oxidase (cyt c oxidase). The reaction of NO with the binuclear metal centre of cyt c oxidase apparently leads to the formation of nitrite at the active site [123] the mechanism of which was described as the opposite of nitrite reduction to NO by non-haem nitrite reductases [124]. The inhibition was caused by the binding of NO to the reduced copper centre of the enzyme rather than the expected reaction with Fe. ... 

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