Molybdopterin-containing enzymes nitrate reductase

Formate dehydrogenases from many bacteria contain molybdopterin and also often selenium (Table 15-4).664/665 A membrane-bound Mo-containing formate dehydrogenase is produced by E. coli grown anaerobically in the presence of nitrate. Under these circumstances it is coupled to nitrate reductase via an electron-transport chain in the membranes which permits oxidation of formate by nitrate (Eq. 18-26). This enzyme is also a multisubunit protein.665 666 Two other Mo- and Se- containing formate dehydrogenases are produced... 

Bacterial assimilatory nitrate reductases have similar properties.86/86a In addition, many bacteria, including E. coli, are able to use nitrate ions as an oxidant for nitrate respiration under anaerobic conditions (Chapter 18). Tire dissimilatory nitrate reductases involved also contain molybdenum as well as Fe-S centers.85 Tire E. coli enzyme receives electrons from reduced quinones in the plasma membrane, passing them through cytochrome b, Fe-S centers, and molybdopterin to nitrate. The three-subunit aPy enzyme contains cytochrome b in one subunit, an Fe3S4 center as well as three Fe4S4 clusters in another, and the molybdenum cofactor in the third.87 Nitrate reduction to nitrite is also on the pathway of denitrification, which can lead to release of nitrogen as NO, NzO, and N2 by the action of dissimi-latory nitrite reductases. These enzymes873 have been discussed in Chapters 16 and 18. 

In all photoautotrophs, reduction of NOj" to NH4 is achieved in two distinct enzymatic steps (Campbell, 2001). First, assimilatory nitrate reductase (NR) catalyzes the two electron reduction from NOj" to NO2. NR is a large soluble cytoplasmic enzyme with FAD (flavin adinine dinucleotide), an iron-containing cytochrome and molybdopterin prosthetic groups, and requires NADH and/or NADPH as an electron donor (Guerrero et al, 1981). Functional NR is in the form of a homodimer and therefore requires two atoms of iron per enzyme. Following transport into the chloroplast, NO2 undergoes a 6 e reduction to NH4 via assimilatory nitrite reductase (NiR). NiR, a soluble chloroplastic enzyme, contains five iron atoms per active enzyme molecule, and requires photosynthetically reduced ferredoxin as an electron donor (Guerrero et al., 1981). 

In the heterotrophic nitrate respiration, in most cases hydrogen atoms derived from organic compounds ([H], mostly in the form of NADH in the case of heterotrophic denitriliers) are first oxidized with nitrate. The reaction is catalyzed by nitrate reductase. The enzyme contains Mo, [Fe4S4] and [Fe2S2] clusters (Fe/S), and cytochrome b (Chaudhry and MacGregor, 1983). Mo is present in the enzyme as molybdenum cofactors combining with molybdopterin or molybdopterin guanine dinucleotide. The enzyme catalyzes the reduction of nitrate to nitrite with ubiquinol or menaquinol (QH2) as the electron donor. 

The sulfite oxidase enzymes are widespread in Nature, and are found in plants, bacteria (the sulfite dehydrogenases) and in birds and mammals. In addition, this family also includes the assimilatory plant nitrate reductases, which have essentially similar molybdenum coordination and differ structurally in lacking an active site arginine that is present in sulfite oxidase, and in showing somewhat different active site structures on turnover. We will focus here on the animal sulfite oxidase enzymes, of which chicken and human are the best studied. In animals the enzyme is responsible for the physiologically essential oxidation of sulfite to sulfate. It is a dimer of 52 kDa subunits and resides in the mitochondrial inner-membrane space. Each monomer contains Mo associated with one molybdopterin, plus a cytochrome heme. The enzymes catalyze the following reaction, which occurs at the Mo site which is reduced from Mo(vi) to Mo(iv) in the process ... 

In the literature, several enzymatic (Kirstein et al., 1999 Cosnier et al., 1994) and nonenzymatic (Gamboa et al., 2009 Groot and Koper, 2004) electrochemical biosensors were reported for the N03 determination. The enzymatic determination of N03 using nitrate reductase (NaR)—modified electrode is novel and highly selective. NaR is a multidomain enzyme containing flavin adenine dinucleotide (FAD), two heme-Fe and molybdopterin, which catalyzes the two-electron reduction of N03 to NOa (Quan et al., 2005). 

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