Monodehydroascorbate reductase

Two ascorbate radicals can react with each other in a disproportionation reaction to give ascorbate plus dehydroascorbate. However, most cells can reduce the radicals more directly. In many plants this is accomplished by NADH + H+ using a flavoprotein monodehydroascorbate reductase.0 Animal cells may also utilize NADH or may reduce dehydroascorbate with reduced glutathione.CC/ff Plant cells also contain a very active blue copper ascorbate oxidase (Chapter 16, Section D,5), which catalyzes the opposite reaction, formation of dehydroascorbate. A heme ascorbate oxidase has been purified from a fungus. 11 1 Action of these enzymes initiates an oxidative degradation of ascorbate, perhaps through the pathway of Fig. 20-2. 

As shown in Figure 13.3, oxidation of ascorbic acid, for example, by the reduction of superoxide to hydrogen peroxide or Fe + to Fe +, and similar reduction of other transition metal ions, proceeds by a one-electron process, forming the monodehydroascorbate radical. The radical rapidly disproportionates into ascorbate and dehydroascorbate. Most tissues also have both nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione-dependent monodehydroascorbate reductases, which reduce the radical back to ascorbate. Ascorbate is thus an effective quencher of singlet oxygen and other radicals. 

Figure 13.3. Redox reactions of ascorbate. Monodehydroascorbate reductase, EC 1.8.5.3 NADPH-dependent dehydroascorbate reductase, EC 1.6.5.4 and glutathione-dependent dehydroascorbate reductase, EC 1.8.5.1.

The immediate product of the oxidative reaction, the monodehydroascorbate radical (Eq. [1]), is a fairly reactive and unstable species which, in the presence of a suitable reductase system, is reduced back to ascorbate. Monodehydroascorbate reductases have been identified and purified in a few cases (Ushimara et al., 1997 Dalton et al., 1992 Shigeoka et al., 1987 Borraccino et al., 1986 Hossain et al., 1984) and cDNA sequences have been published for the pea (Murthy and Zilinskas, 1994) and cucumber enzymes (Sano et al., 1995). A bacterial expression system is also available for the cucumber enzyme (Sano et al., 1995). In the absence of a suitably efficient reductase system, the monodehydroascorbate radicals disproportionate to dehydroascorbate and ascorbate in this case, ascorbate is regenerated using a glutathione-dependent dehydroascorbate reductase enzyme (Foyer and Mullineaux, 1998 and references therein). Under non-... 

Hossain, M. A., Nakano, Y., and Asada, K., 1984, Monodehydroascorbate reductase in spinach chloroplasts and its participation in regeneration of ascorbate for scavenging hydrogen peroxide. Plant Cell Physiol. 25 3859395. 

Murthy, S. S., and Zilinskas, B. A., 1994, Molecular cloning and characterisation of a cDNA encoding pea monodehydroascorbate reductase, J. Biol. Chem. 269 31129n 31133. 

Ushimara, T., Maki, Y., Sano, S., Koshiba, K., Asada, K., and Tsuji, H., 1997, Induction of enzymes involved in the ascorbate-dependent antioxidative systems, namely ascorbate peroxidase, monodehydroascorbate reductase and dehydroascorbate reductase, after exposure to air of rice Oryza sativa) seedlings germinated under water, Plant Cell Physiol. 38 541n549. 

Fig. 14. The ascorbate-glutathione cycle. APX - ascorbate peroxidase DMA - dehydroascorbate DHAR - dehydroascorbate reductase OR - glutthione reductase GSH - reduced glutathione GSSG - glutathione disulfide MDHR - monodehydroascorbate reductase.

Pulse radiolysis was also used to elucidate the mechanism of catalytic action of monodehydroascorbate reductase, an enzyme containing FAD and using Nicotinaminde atjenine dinucleotide (NADH) as reductant. The substrate is dehydroascorbate radical produced by pulse radiolysis (130). The authors show that this radical reacts with the protein to give the FADH radical and that the... 

Hou, W.-C. and Lin, Y-H. 1997. Dehydroascorbate reductase and monodehydroascorbate reductase activities of trypsin inhibitors, the major sweetpotato (Ipomoea batatas [L.] Lam) root storage protein. Plant Sci. 149, 151-156. 

Enzyme assays Intact chloroplasts were isolated and then osmotic-ally shocked. The lysates were used for determination of the stromal activities of ascorbate peroxidase (6), monodehydroascorbate reductase (7) and dehydroascorbate reductase (6). Superoxide dismutase (SOD) was assayed (8) in a protein extract derived from chloroplasts solubilized in the presence of 2.5% Triton-X-100. Catalase activity was determined (9) in leaf homogenates. 

Goldenberg, H., Grebing, C., and Low, H., 1983, NADH-monodehydroascorbate reductase in human erythrocyte plasma membrane, Biochem. Int. 6 1-10. 

Schweinzer, E., and Goldenberg, H., 1993, Monodehydroascorbate reductase activity in the surface membrane of leukemic cells. Characterization by a ferricyanide-driven redox cycle, Eur. J. Bio-chem. 218 1057-1062. 

Oxidoreductases (such as ascorbate oxidase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, superoxide dismutase and ascorbate cytochrome-fo reductase) that act in the metabolism of vitamin C in animals and plants can be considered to be antivitamins C. Other oxidoreductases, such as enzymes known trivially as polyphenoloxidases and some others, may indirectly cause loss of ascorbic acid. 

As shown in Figure 1, oxidation of ascorbic acid proceeds by a one-electron process, forming mono-dehydroascorbate, which disproportionates to ascorbate and dehydroascorbate. Most tissues also contain monodehydroascorbate reductase (EC 1.6.5.4), a flavoprotein that reduces the radical back to ascorbate. Dehydroascorbate is reduced to ascorbate by dehydroascorbate reductase (EC 1.8.5.1), a glutathione-dependent enzyme little is oxidized to diketogulonic acid in human beings. 

Reactive oxygen species (ROS) and other radicals are effectively quenched by carotenoids, tocopherols, ascorbate, and reduced glutathione, supported by antioxidant enzymes, including superoxide dismutase (SOD), catalase, glutathione peroxidase (GSH-Px), and the enzymes involved in the ascorbate-glutathione cycle to detoxify ROS, such as ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR). " ... 

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