NADH and NADPH

Nicotinamide is an essential part of two important coenzymes nicotinamide adenine dinucleotide (NAD ) and nicotinamide adenine dinucleotide phosphate (NADP ) (Figure 18.19). The reduced forms of these coenzymes are NADH and NADPH. The nieotinamide eoenzymes (also known as pyridine nucleotides) are electron carriers. They play vital roles in a variety of enzyme-catalyzed oxidation-reduction reactions. (NAD is an electron acceptor in oxidative (catabolic) pathways and NADPH is an electron donor in reductive (biosynthetic) pathways.) These reactions involve direct transfer of hydride anion either to NAD(P) or from NAD(P)H. The enzymes that facilitate such... 

Synthesis and reactions of NADH and NADPH model compounds with interconversion of central and axial chirality 97YGK132. 

NAD+ and NADP+ are coenzymes of dehydrogenases. NADH and NADPH are intermediate carriers of both hydrogen and electrons. Most NAD-dependent enzymes are located in the mitochondria and deliver H2 to the respiratory chain whereas NADP-dependent enzymes take part in cytosolic syntheses (reductive biosyntheses). 

The significance of these metabolites in the biosynthesis of the thiamine thiazole in considered next. Although, from their constitution, and from the tracer experiments, the metabolites are undoubtedly the products of transformation of 1-deoxy-D-t/ireo-pentulose, their significance in the biosynthesis of the thiazole of thiamine is not clear. The thiazole glycol is not a product arising from a transformation of the thiazole (5) of thiamine. Reduction to this thiazole (5) occurs in dialyzed extracts of disrupted cells, in the presence of ATP, NADH, and NADPH, but only at 0.2% the rate of synthesis of the thiamine thiazole (5) by intact cells. The behavior of the thiazole glycol on plates is merely a consequence of the extreme sensitivity of the tetrazolium reagent. 

The physicochemical properties of the reactants in an eiKyme-catalyzed reaction dictate the options for the assay of enzyme activity. Spectrophotometric assays exploit the abihty of a substrate or product to absorb hght. The reduced coenzymes NADH and NADPH, written as NAD(P)H, absorb hght at a wavelength of 340 run, whereas their oxidized forms NAD(P) do not (Figure 7—9). When NAD(P)+ is reduced, the absorbance at 340 run therefore increases in proportion to—and at a rate determined by—the quantity of NAD(P)H produced. Conversely, for a dehydrogenase that catalyzes the oxidation of NAD(P)H, a decrease in absorbance at 340 run will be observed. In each case, the rate of change in optical density at 340 nm will be proportionate to the quantity of enzyme present. 

In addition to moving acetyl-CoA from the mitochondria to the cytoplasm, this cycle also converts an NADH to an NADPH. If we assume that the amount of ATP that we could get from NADH and NADPH oxidation is the same, making NADPH from NADH and NADP+ doesn t cost any energy. So we can conclude that the cost of just moving the acetyl-CoA out of the mitochondria is 2 ATPs per acetyl-CoA. 

Peroxynitrite reacts with heme proteins such as prostacycline synthase (PGI2), microperoxidase, and the heme thiolate protein P450 to form a ferryl nitrogen dioxide complex as an intermediate [120]. Peroxynitrite also reacts with acetaldehyde with the rate constant of 680 1 mol 1 s" 1 forming a hypothetical adduct, which is decomposed into acetate, formate, and methyl radicals [121]. The oxidation of NADH and NADPH by peroxynitrite most certainly occurs by free radical mechanism [122,123], Kirsch and de Groot [122] concluded that peroxynitrite oxidized NADH by a one-electron transfer mechanism to form NAD and superoxide ... 

Similar to LOXs, cyclooxygenases may catalyze superoxide production in the presence of NADH and NADPH [49]. It has been shown [88] that prostaglandin H synthase produced oxygen radicals and hydrogen peroxide during the transformation of 2(3)-tcrt-butyl-4-... 

The exact nature of the NO-releasing reaction and the other products of reaction in mammalian tissue are still unclear. The matter has been discussed by a number of authors and a reductive mechanism in rat hepatocytes and human erythrocytes has been suggested in the presence of NADH and NADPH. Nitroprusside can pass through cell membranes and so there is no intrinsic difficulty with this suggestion. There is direct evidence from spin echo NMR studies to show the conversion, by nitroprusside, of glutathione into glutathione disulfide within erythrocytes [49]. 

Numerous experiments were then performed with mitochondria incubated with acetate, CoA, ATP, etc., in attempts to detect fatty acid synthesis. In 1957, Lynen and his colleagues reported the presence in mitochondria of a system which catalyzed the elongation of caproyl CoA to octanoyl CoA by the addition of an acetate unit. NADH and NADPH had to be present. The existence of this mitochondrial system was confirmed by Wakil et al. in 1961 who showed the 12C acid could be extended to 16C by successive additions of 2C fragments. 

Intracellular reduced pyridine nucleotides NAD(P)H are the primary suppliers of reducing power to anabolic and catabolic pathways. They can be measured because of their fluorescent properties.<16) The fluorescence is caused by the presence of the reduced forms of the pyridine nucleotides NADH and NADPH (jointly referred to as NAD(P)H). These fluorophores absorb light in a wide band around 340 nm, and reemit, or fluoresce, light in a wide band around 460 nm. The phosphorylated and nonphos-phorylated nucleotides have essentially equivalent fluorescence properties while the oxidized forms of these nucleotides are nonfluorescent. 

Tab. 13.6 Overview of retentions of various membranes for NAD, NADP, NADH and NADPH in different solutions.

The reverse reaction, i.e. reduction, is also indicated in the scheme, and may be compared with the chemical reduction process using complex metal hydrides, e.g. LiAlPLj or NaBH4, namely nucleophilic addition of hydride and subsequent protonation (see Section 7.5). The reduced forms NADH and NADPH are conveniently regarded as hydride-donating reducing agents (see Box 7.6). We also noted that there were stereochemical features associated with these coenzymes (see Box 3.14). During a reduction... 

We should note that there are other intrinsic fluoro-phores that can be used in binding studies. NADH and NADPH are both highly fluorescent coenzymes. So too are riboflavin and FAD, but flavoproteins are not typi-... 

I. 14.13.33] can utilize NADH and NADPH equally well. The enzyme isolated from Corynebacterium cyclohexani-cum is highly specific for 4-hydroxybenzoate. 

The oxidation of 1,2- and 1,4-dihydropyridines has been extensively studied. This is due in large part to the occurrence of the 1,4-dihydropyridine ring system in the reduced forms of the coenzymes nicotinamide adenine di- and tri-phosphate (NADH and NADPH). These redox couples are responsible for a number of biological oxidations and reductions (B-70MI20701). 

Depletion of other cofactors such as UTP, NADH, and NADPH may also be involved in cell injury either directly or indirectly. Thus, the role of NADPH in maintaining reduced GSH levels means that excessive GSH oxidation such as caused by certain quinines, which undergo redox cycling, may in turn cause NADPH depletion (see below). Alternatively, NADPH may be oxidized if it donates electrons to the foreign compound directly. However, NADPH may be regenerated by inter conversion of NAD+ to NADP+. Some quinones such as menadione, l,2-dibromo-3-chloropropane (DBCP), and hydrogen peroxide also cause depletion of NAD, but probably by different mechanisms. Thus, with menadione, the depletion may be the result of... 

NADH and NADPH Act with Dehydrogenases as Soluble Electron Carriers... 

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