Nitrate reductase, denitrifying bacteria

Nitrite reductases (NiRs)—enzymes found in several strains of denitrifying bacteria— catalyze the one-electron reduction of nitrite anion to nitric oxide (Equation 1). - In addition to the importance of this process in the global nitrogen cycle (Figure 1), further incentive for the study of the denitrification process is provided by its environmental impact, ranging from the production of NO as a pollutant and NjO as a potent greenhouse gas, to lake eutrophication due to farm runoff that contains high concentrations of nitrates and nitrites. 

Calmels, S., Ohshima, H., and Bartsch, H. (1988). Nitrosamine formation by denitrifying and non-denitryfying bacteria Implications of nitrite reductase and nitrate reductase in nitrosation catalysis. . Gen. Microbiol. 134, 221-226. 

A number of other reductases and dehydrogenases, including dissimilatory nitrate reductases of E. coli and of denitrifying bacteria (Chapter 18), belong to the DMSO reductase family. Other members are reductases for biotin S-oxide,649 trimethylamine N-oxide, and polysulfides as well as formate dehydrogenases (Eq. 16-63), formylmethanofuran dehydrogenase (Fig. 15-22,... 

NapB. NapB is a subunit of the heterodimeric periplasmic nitrate reductase (NapAB) and transfers electrons to the catalytic NapA molybdoprotein. Nap systems are found in a number of bacteria, iucluding enterobacteria, aerobic denitrifiers, and nonsulfur purple photo synthetic bacteria. Their physiological function is different in these groups of bacteria and includes redox balancing using nitrate as an electron sink to dispose of excess rednctant, aerobic denitrification, and nitrate scavenging in nitrate-limited environments. 

Nitrate can also be converted to N02 using cadmium reduction (columns or spongy cadmium) as described above. Once the NOa" is in the form of N02, the N02 can be isolated via organic extraction (e.g., Olson, 1981) or with SPE after the N02 is converted to an azo dye (Kator et al, 1992). Nitrate can be isolated by conversion to N2O via sodium azide in an acetic acid buffer solution (Mcllvin and Altabet, 2005). Another approaches uses a genetically engineered denitrifier to convert N03 to N2O (Sigman et al, 2001) the bacteria wiU denitrify NOa" in a sample to N2O, but lacking nitrous oxide reductase the bacteria cannot take the reaction to completion and form N2. The N2O produced by either approach can then be analyzed on a mass spectrometer. A more detailed discussion of these methods is presented in Chapter 31 by Lipschultz, this volume. 

Nitrate and NO2 reduction are rather widespread in SO reducers (Keith and Herbert, 1983 McCready et al., 1983 Mitchell et al., 1986 Maumetal., 1997 Seitz and Cypionka, 1986), and in some cases NOj" is preferred over SO (Seitz and Cypionka, 1986). The bisulfite reductase enzyme in SO reducers displays some activity toward NO2, which partially explains the ubiquity of NO2 reduction. However, it appears that specific NO2 reductases are also present (Liu and Peck Jr., 1981). Unlike denitrifying bacteria that reduce NOj" to N2, the end product of NOj" reduction in SO reducers is NH4 (Widdel and... 

Denitrifying bacteria generally contain cytochrome hi as the electron donor to the nitrate reductase (197,198). This cytochrome has been partially purified from M. dentrificans (163,164), and this preparation (absorption maxima at 559, 528, and 426 nm) was rapidly oxidized by nitrate. A similar cytochrome, 6-558, was solubilized from pseudomonads (163,164,199). Another similar cytochrome, h-559, is contained, together... 

Two types of dissimilatory nitrate reductase (NR) have been characterized in denitrifying bacteria, being either membrane-bound or soluble enzymes. Nitrate reduction by membrane-bound NR (encoded by nar) occurs in the cytoplasm and is involved in energy conservation, in contrast... 

The denitrification process could be described as a modular organization in which every biochemical reaction is catalyzed by specific reductase enzymes (Cuervo-Lopez et al., 2009). Four enzymatic reactions take place in the cell as follows (l) nitrate is reduced to nitrite by nitrate reductase (Nar) (ii) a subsequent reduction of nitrite to nitric oxide is carried out by nitrite reductase (Mr) (iu) afterwards, nitric oxide is reduced to nitrous oxide by the enzyme nitric oxide reductase (Nor) (iz ) finally, nitrous oxide is reduced to N2 by the enzyme nitrous oxide reductase (Nos) (Lalucat et al., 2006) (Table 9). These reactions take place when environmental conditions become anaerobic (Berks et al., 1995 Hochstein Tomlinson, 1988). The enzymatic reactions, which are thermodynamically favored, are carried out in the cell membrane and periplasmic space. Small half saturation constant values (Km) have been reported for different nitrogen substrates for some denitrifying bacteria, indicating that denitrifying enzymes have a high affinity for their substrate. However, several factors have to be considered, as the presence of small quantities of molybdenum, cooper and hem to ensure the successful enzymatic activity, as they are known cofactors for denitrifying enzymes. 

It was long believed that bacteria were unique in their ability to denitrify. However, Shoun and Tanimoto (1991) and Shoun et al., (1989) demonstrated that the fungus, Fusarium oxysporum, could be induced to synthesize an enzyme system capable of the anaerobic reduction of nitrite to N2O. Induction occurred under conditions of low oxygen concentrations in the presence of nitrate or nitrite. One and pethaps the only component of this nitrite reductase system is a unique, soluble cytochrome P-450 (P-450dNIR), which is more similar in its cDNA-inferred amino acid sequence to soluble, bacterial P-450 enzymes (espe-... 

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