Flavin adenine dinucleotide reductase reduction

Nitrobenzene reductase activity has been detected in the fat body, gut, and Malpighian tubules of the Madagascar cockroach, G. portentosa (Rose and Young, 1973). Anaerobic conditions are essential for activity. The enzymes in the microsomes are strongly NADH dependent, whereas those in the soluble fraction are strongly NADPH dependent. Activity is enhanced by the addition of flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN) or riboflavin. It appears that the true substrate for the nitroreductase is FMN and that the reduction of the nitro compounds occurs nonenzymatically (Figure 8.15). Similar results are obtained using azofuchsin as substrate. 

Methylene-Tetrahydrofolate Reductase The reduction of methylene-tetrahydrofolate to methyl-tetrahydrofolate, shown in Figure 10.7, is catalyzed hy methylene-tetrahydrofolate reductase, a flavin adenine dinucleotide-dependent enzyme during the reaction, the pteridine ring of the substrate is oxidized to dihydrofolate, then reduced to tetrahydrofolate by the flavin, which is reduced by nicotinamide adenine dinucleotide phosphate (NADPH Matthews and Daubner, 1982). The reaction is irreversible under physiological conditions, and methyl-tetrahydrofolate - which is the main form of folate taken up into tissues (Section 10.2.2) - can only be utilized after demethylation catalyzed by methionine synthetase (Section 10.3.4). 

Bacterial mercuric reductase is a unique metal-detoxification biocatalyst, reducing mercury(II) salts to the metal. The enzyme contains flavin adenine dinucleotide, a reducible active site disulfide (Cys 135, Cys i4o), and a C-terminal pair of cysteines (Cys 553, Cys 559). Mutagenesis studies have shown that all four cysteines are required for efficient mercury(II) reduction. Mercury Lm-EXAFS studies for mercury(II) bound to both the wild-type enzyme and a very low-activity C-terminal double-alanine mutant (Cys 135, Cys uo, Ala 553, Ala 559) suggest the formation of an Hg(Cys)2 complex in each case (39). The Hg—S distances obtained were 2.31 A and are consistent with the correlation of bond length with coordination number presented above. Thus, no evidence was obtained for coordination of mercury(II) by all four active-site cysteines in the wild-type mercuric reductase. However, these studies do not define the full extent of the catalytic mechanism for mercury(II) reduction, and it is possible that a three- or four-coordinate Hg(Cys) complex is a key intermediate in the process. 

Glutathione reductase (EC 1.8.1.7 formerly EC 1.6.4.2 GSR) links the glutathione pathway to the hexose monophosphate pathway through the reversible oxidation and reduction of NADP. Flavin adenine dinucleotide (FAD) is necessary as a cofactor. The enzyme maintains high levels of reduced glutathione in the cell. Two isoforms exist, a mitochondrial and cytoplasmic form, produced by alternative initiation. The molecule is a homodimer, linked by a disulfide bridge. Each subunit (522 amino acids 56kDa) is divided into four domains of which domains one and two bind FAD and NADPH, respectively and domain four forms the interface. ... 

FIGURE 2 Proposed dual mode for calmodulin (CaM) control of nitric oxide synthase (NOS) electron transfer. Neuronal NOS is composed of a reductase and an oxygenase domain, shown as two circles. CaM binding to NOS activates at two points in the electron transfer sequence (1) It increases the rate at which NADPH-derived electrons are transferred into the flavins, and (2) it enables the flavins to pass electrons to the oxygenase domain of NOS. Activation at the first point is associated with an increase in reductase domain-specific catalytic activities, such as electron transfer to cytochrome c or ferricyanide (FeCN ). Activation at the second point is associated with a reduction of NOS heme iron, an initiation of NO synthesis from L-arginine (Arg), or a reduction of Oj to form superoxide (O2) in the absence of substrate. FAD, Flavin-adenine dinucleotide FMN, flavin mononucleotide NO, nitric oxide. 

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). 

Plants usually obtain their nitrogen by absorption of nitrate or ammonium ions from the soil (symbiotic associations between higher plants and nitrogen fixing bacteria are of course exceptions to this). Ammonium ions may be utilized directly in the synthesis of amino acids (see p. 169), but nitrate must first be reduced to ammonia. This is accomplished in two stages the reduction of nitrate to nitrite followed by the reduction of nitrite to ammonia. The first step— nitrate reduction—is catalysed by the flavo-protein enzyme complex nitrate reductase (Fig. 5.12) which contains molybdenum and FAD (flavin adenine dinucleotide) as a prosthetic group. Reduced FMN... 

The nitrite reductase of Azotobacter vinelandvi (A. agile) was extracted in soluble form by Nason and his colleagues (537). The preparation reduced nitrite and hydroxylamine in the presence of reduced nicotinamide-adenine dinucleotides and required flavin for maximal activity. FAD was shown to be specific for nitrite reduction, whereas both FAD and FMN were active for hydroxylamine reduction. The hydroxylamine reductase activity of the preparation was enhanced in the presence of Mn +. Ammonia was shown to be the product of nitrite reduction, but the product of hydroxylamine reduction was not identified. Another nitrite and hydroxylamine reductase, which had a Mn requirement, was also isolated and partially purified in Nason s laboratory from soybean... 

Dehydrogenases and reductases require a coenz5mie such as nicotinamide adenine dinucleotide (NADH), nicotinamide adenine dinucleotide phosphate (NADPH), flavin, etc. The reduction of a carbonyl compoxmd with NAD(P)H with auxiliary substance proceeds as follows (Figure 11.1a). 

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