NADP Biosynthesis

NAD and NADP are required as redox coen2ymes by a large number of enzymes and ia particular dehydrogenases (Fig. 6). NAD" is utilized ia the catabohe oxidations of carbohydrates, proteins, and fats, whereas NADPH2 is the coenzyme for anaboHc reactions and is used ia fats and steroid biosynthesis. NADP+ is also used ia the cataboHsm of carbohydrates (2). 

NADP. H2 is used to reduce fatty acid biosynthesis intermediates ... 

The conclusion that the significance of the pentose phosphate shunt may be in keeping NADP in its reduced state and furnishing pentose phosphates for biosynthesis, rather than G-6-P utilization, is closely confirmed by the fact that individuals deficient in or lacking G-6-PDH activity suffer from a number of metabolic disorders due to lack of NADPH2 generation and nucleotide depletion. 

Nicotinate and nicotinamide, together referred to as niacin, are required for biosynthesis of the coenzymes nicotinamide adenine dinucleotide (NAD"") and nicotinamide adenine dinucleotide phosphate (NADP" ). These both serve in energy and nutrient metabolism as carriers of hydride ions (see pp. 32, 104). The animal organism is able to convert tryptophan into nicotinate, but only with a poor yield. Vitamin deficiency therefore only occurs when nicotinate, nicotinamide, and tryptophan are all simultaneously are lacking in the diet. It manifests in the form of skin damage (pellagra), digestive disturbances, and depression. 

In hepatocytes and adipocytes, cytosolic NADPH is largely generated by the pentose phosphate pathway (see Fig. 14-21) and by malic enzyme (Fig. 21-9a). The NADP-linked malic enzyme that operates in the carbon-assimilation pathway of C4 plants (see Fig. 20-23) is unrelated in function. The pyruvate produced in the reaction shown in Figure 21-9a reenters the mitochondrion. In hepatocytes and in the mammary gland of lactating animals, the NADPH required for fatty acid biosynthesis is supplied primarily by the pentose phosphate pathway (Fig. 21-9b). 

This unique redox catalyst links the oxidation of H2 or of formate to the reduction of NADP+229 and also serves as the reductant in the final step of methane biosynthesis (see Section E) 228 It resembles NAD+ in having a redox potential of about -0.345 volts and the tendency to be only a two-electron donor. More recently free 8-hydroxy-7,8-didemethyl-5-deazaribo-flavin has been identified as an essential light-absorbing chromophore in DNA photolyase of Methanobacterium, other bacteria, and eukaryotic algae.230 Roseoflavin is not a coenzyme but an antibiotic from Streptomyces davawensisP1 Many synthetic flavins have been used in studies of mechanisms and for NMR232 and other forms of spectroscopy. 

The enzyme is organized into two structural domains,201 one of which binds FAD and the other NADP+. Similar single-electron transfers through flavoproteins also occur in many other enzymes. Chorismate mutase, an important enzyme in biosynthesis of aromatic rings (Chapter 25), contains bound FMN. Its function is unclear but involves formation of a neutral flavin radical.276 277... 

As a general rule, NAD+ is associated with catabolic reactions and it is somewhat unusual to find NADP+ acting as an oxidant. However, in mammals the enzymes of the pentose phosphate pathway are specific for NADP+. The reason is thought to lie in the need of NADPH for biosynthesis (Section I). On this basis, the occurrence of the pentose phosphate pathway in tissues having an unusually active biosynthetic function (liver and mammary gland) is understandable. 

Still another difference between biosynthesis of fatty acids and oxidation (in mammals) is that the former has an absolute requirement for NADPH (Fig. 17-12) while the latter requires NAD+ and flavo-proteins (Fig. 17-1). This fact, together with many other observations, has led to the generalization that biosynthetic reduction reactions usually require NADPH rather than NADH. Many measurements have shown that in the cytosol of eukaryotic cells the ratio [NADPH]/[NADP+] is high, whereas the ratio [NADH]/[NAD+] is low. Thus, the NAD+/NADH system is kept highly oxidized, in line with the role of NAD+ as a principal biochemical oxidant, while the NADP+/NADPH system is kept reduced. 

To understand why isocitrate dehydrogenase is so intensely regulated we must consider reactions beyond the TCA cycle, and indeed beyond the mitochondrion (fig. 13.15). Of the two compounds citrate and isocitrate, only citrate is transported across the barrier imposed by the mitochondrial membrane. Citrate that passes from the mitochondrion to the cytosol plays a major role in biosynthesis, both because of its immediate regulatory properties and because of the chain of covalent reactions it initiates. In the cytosol citrate undergoes a cleavage reaction in which acetyl-CoA is produced. The other cleavage product, oxaloacetate, can be utilized directly in various biosynthetic reactions or it can be converted to malate. The malate so formed can be returned to the mitochondrion, or it can be converted in the cytosol to pyruvate, which also results in the reduction of NADP+ to NADPH. The pyruvate is either utilized directly in biosynthetic processes, or like malate, can return to the mitochondrion. 

Comparing this equation with the equation for the complete oxidation of palmitoyl-CoA (see table 18.1, equation 1), we find major differences in carriers and intermediates. The principal electron carrier in the anabolic pathway is the NADPH-NADP+ system in the catabolic pathway, /3 oxidation, the principal electron carriers are FAD-FADH2 and NAD+-NADH. The second striking difference between the two pathways is that malonyl-CoA is the principal substrate in the anabolic pathway but plays no role in the catabolic pathway. These differences reflect the fact that the two pathways do not share common enzymes. Indeed, in animal cells the reactions occur in separate cell compartments biosynthesis takes place in the cytosol, whereas catabolism occurs in the mitochondria. 

Nucleotides play important roles in all major aspects of metabolism. ATP, an adenine nucleotide, is the major substance used by all organisms for the transfer of chemical energy from energy-yielding reactions to energy-requiring reactions such as biosynthesis. Other nucleotides are activated intermediates in the synthesis of carbohydrates, lipids, proteins, and nucleic acids. Adenine nucleotides are components of many major coenzymes, such as NAD+, NADP+, FAD, and CoA. (See chapter 10 for structures of these coenzymes.)... 

Glucose-6-phosphate dehydrogenase, 6-phosphoglu-conate dehydrogenase, and the NADP+-malic enzyme are sources of the 14 moles of NADPH required for biosynthesis of palmitate. 

The reduction of />coumaroyl-CoA (3.31) to />coumaryl aldehyde (3.69) is catalyzed by the enzyme cinnamoyl-CoA NADP oxidoreductase (CCR). This enzyme was initially purified from soybean cultures (Wegenmayer et al., 1976), and was later on efficiently isolated from lignifying cambium of eucalyps (Eucalyptus gunnii) (Gofifiier et al., 1994). A CCR cDNA was identified in a cDNA library that was screened with oligonucleotiede derived from the peptide sequence of the CCR protein. CCR is considered the first enzyme committed towards the biosynthesis of monolignols and shows... 

Further evidence32 has been obtained which strongly supports the conclusion (c/. Vol. 10, p. 19) that geissoschizine (101) is at a shunt in the biosynthesis of the three tissue-culture alkaloids (102)—(104). Incubation of (101) with the enzyme preparation plus NADP+ and NADPH in D20 afforded a sample of (103) that contained one deuterium atom per molecule. Geissoschizine (101) must thus be involved in biosynthesis after (98). With NADPD as co-factor, a sample of (103) was isolated that contained a single deuterium atom, located at C-21, in each molecule. Moreover, geissoschizine with deuterium that was stereochemically a at C-21 gave (103) that was devoid of label. These two results support biosynthesis via (99). 

Two distinct IDHs have been identified in living organisms NAD-IDH (EC 1.1.1.41) is limited exclusively to eukaryotic organisms, and NADP-IDH (EC 1.1.1.42) is ubiquitously present in both prokaryotes and eukaryotes [15], These enzymes play important biological roles NAD-IDH is involved in the supply of NADH used for respiratory ATP production in mitochondria of eukaryotic organisms the bacterial NADP-IDH is also part of the Krebs cycle machinery. However, the eukaryotic NADP-dependent enzyme is primarily involved in providing NADPH and a-ketoglutarate for biosynthesis [15],... 

Reactions (10.9) and (10.10) are catalyzed by flavokinase and FMN pyrophos-phorylase, respectively. For the structures of FMN and FAD, see Chapter 6. The biosynthesis of NAD+ from nicotinamide is shown in Figure 10.12. An additional phosphorylation step with ATP is required to form NADP+. If one... 

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