Nitrate reductase electron, donors

Similar mechanisms operate in the action of nitrate reductase and nitrite reductase. Both of these substances are produced from ammonia by oxidation. Plants and soil bacteria can reduce these compounds to provide ammonia for metabolism. The common agricultural fertilizer ammonium nitrate, NH4NO3, provides reduced nitrogen for plant growth directly, and by providing a substrate for nitrate reduction. NADH or NADPH is the electron donor for nitrate reductase, depending on the organism. 

Many species of bacteria also have an assimilatory nitrite reductase which is located in the cytoplasm. There is relatively little known about such enzymes but the electron donor is throught to be NADPH and the active site again has siroheme (Cole, 1988). The assimilatory nitrite reductases of both plants and bacteria use nitrite that is provided as the product of the assimilatory nitrate reductases. Nitrate is a very common natural N source for plant and bacterial growth. 

The [2Fe 2S], [3Fe S], and [4Fe S] clusters that are found in simple Fe S proteins are also constituents of respiratory and photosynthetic electron transport chains. Multicluster Fe S enzymes such as hydrogenase, formate dehydrogenase, NADH dehydrogenase, and succinate dehydrogenase feed electrons into respiratory chains, while others such as nitrate reductase, fhmarate reductase, DMSO reductase, and HDR catalyze the terminal step in anaerobic electron transport chains that utihze nitrate, fumarate, DMSO, and the CoB S S CoM heterodisulfide as the respiratory oxidant. All comprise membrane anchor polypeptide(s) and soluble subunits on the membrane surface that mediate electron transfer to or from Mo cofactor (Moco), NiFe, Fe-S cluster or flavin active sites. Multiple Fe-S clusters define electron transport pathways between the active site and the electron donor or... 

By the second approach, the enzyme is immobilized in a redox polymer assembly (Figure 39B). Electron-transfer quenching of the photosensitizer by the polymer matrix generates an electron pool for the activation of the enzyme. Photoreduction of nitrate to nitrite was accomplished by the physical encapsulation of NitraR in a redox-functionalized 4,4 -bipyridinium acrylamide copolymer [234]. In this photosystem, Ru(bpy)3 + was used as a photosensitizer and EDTA as a sacrificial electron donor. Oxidation of the excited photosensitizer results in electron transfer to the redox polymer, and the redox sites on the polymer mediate further electron transfer to the enzyme redox center, where the biocatalyzed transformation occurs. The rate constant for the MET from the redox polymer functionalities to the enzyme active site is — (9 + 3) x 10 s. Similarly, the enzyme glutathione reductase was electrically wired by interacting the enzyme with a redox polymer composed of polylysine modified with A-methyl-A -carboxyalkyl-4,4 -bipyridinium. The photosensitized reduction of oxidized glutathione (GSSG) (Eq. 21) ... 

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). 

This two-electron reduction uses NADH or NADPH as electron donor depending on the particular nitrate reductase (EC 1.7.1.1 or EC 1.7.1.2, respectively). Nitrite is then reduced to ammonium in a six-electron process that involves the transfer of three electron pairs from NAD(P)H ... 

Although conducting polymers have demonstrated direct electrochemical communication with nitrate reductases, the incorporation of electron relay groups within the polymer matrix provides a more efficient pathway for electron hopping between the enzyme and the electrode surface. Several artificial electron donors can shuttle electrons to the oxidized form of nitrate reductase with methyl viologen being the best choice due to its very negative redox potential [214-216]. The electron transfer reactions can be represented as follows ... 

The two-domain, structural motif in FNR represents a common structural feature in a large class of enzymes that catalyze electron transfer between a nicotinamide dinucleotide molecule and a one-electron carrier. Beside the photosynthetic electron-transfer enzyme, others non-photosynthetic ones include flavodoxin reductase, sulfite reductase, nitrate reductase, cytochrome reductase, and NADPH-cyto-chrome P450 reductase. FNR belongs to the group of so-called dehydrogenases-electron transferases, i.e., flavoproteins that catalyze electron transfer from two, one-electron donor molecules to a single two-electron acceptor molecule. 

The periplasmatic vanadium-containing nitrate reductase from P. isachenkovii has a molecular mass of 220 kDa (four subunits). The pterin cofactor is again absent. In media supplemented with vanadate and nitrate, vanadate is first reduced by a membrane-bound reductase using NADH as electron donor. This dissimilatory reduction was followed by nitrate consumption.I " ]... 

Fig. 3.7. A schematic presentation of the reaction processes in the nitrate respiration. [H], hydrogen atom bound to certain compounds Q, ubiquinone Cyt, cytochrome. Downward arrow means that ATP is biosynthesized using the energy released by the corresponding electron transfer systems. Nitric oxide reductase is also known to use quinol as the electron donor (see text and Chapter 2, p. 13)...

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. 

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... 

The last three steps in the reduction of nitrate to ammonia are carried out by an enzyme called nitrite reductase. It contains one Fe2S2 center and one molecule of siroheme, a partially reduced iron porphyrin. The electron donor for each step is ferredoxin. 

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