title: Monodehydroascorbate reductase from cucumber is a flavin adenine dinucleotide enzyme.
abstract: Monodehydroascorbate reductase (EC 1.6.5.4) was purified from cucumber fruit to a homogeneous state as judged by polyacrylamide gel electrophoresis. The cucumber monodehydroascorbate reductase was a monomer with a molecular weight of 47,000. It contained 1 mol of FAD/mol of enzyme which was reduced by NAD(P)H and reoxidized by monodehydroascorbate. The enzyme had an exposed thiol group whose blockage with thiol reagents inhibited the electron transfer from NAD(P)H to the enzyme FAD. Both NADH and NADPH served as electron donors with Km values of 4.6 and 23 microM, respectively, and Vmax of 200 mol of NADH and 150 mol of NADPH oxidized mol of enzyme-1 s-1. The Km for monodehydroascorbate was 1.4 microM. The amino acid composition of the enzyme is presented. In addition to monodehydroascorbate, the enzyme catalyzed the reduction of ferricyanide and 2,6-dichloroindophenol but showed little reactivity with calf liver cytochrome b5 and horse heart cytochrome c. The kinetic data suggested a ping-pong mechanism for the monodehydroascorbate reductase-catalyzed reaction. Cucumber monodehydroascorbate reductase occurs in soluble form and can be distinguished from NADPH dehydrogenase, NADH dehydrogenase, DT diaphorase, microsome-bound NADH-cytochrome b5 reductase, and NADPH-cytochrome c reductase by its molecular weight, amino acid composition, and specificity of electron acceptors and donors.
mesh_terms: D000596:Amino Acids / Q000032:analysis; D003683:Dehydroascorbic Acid / Q000378:metabolism; D004355:Drug Stability; D005182:Flavin-Adenine Dinucleotide / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010944:Plants / Q000201:enzymology*; D013053:Spectrophotometry; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D009243:NAD; D005182:Flavin-Adenine Dinucleotide; D009249:NADP; D010088:Oxidoreductases; C020666:glutathione dehydrogenase (ascorbate); D003683:Dehydroascorbic Acid