title: Purification and properties of L-galactono-gamma-lactone dehydrogenase, a key enzyme for ascorbic acid biosynthesis, from sweet potato roots.
abstract: L-Galactono-gamma-lactone dehydrogenase (L-galactono-gamma-lactone:ferricytochrome c oxidoreductase [EC 1.3.2.3], GLDHase) which catalyzes the terminal step in the biosynthesis of L-ascorbic acid (AsA) has been purified from roots of sweet potato (Ipomoea batatas L., cv. Kintoki). Highly purified preparation of the GLDHase was obtained by three column chromatography steps with a recovery of ca. 1%, after solubilization from mitochondria in sweet potato roots. SDS-PAGE exhibited a single band at 56 kDa. In the native state, the apparent molecular mass of the enzyme was 56 kDa, based on a Sephadex G-100 gel filtration. The pI and optimum pH values were 5.8 and 7.9, respectively. The Km value for L-galactono-gamma-lactone was 0.12 mM. Substrate inhibition was obtained at concentrations greater than 4.2 mM. The enzyme was inhibited by p-chloromercuribenzoate (PCMB) and acriflavine, and the inhibition of acriflavine was diminished by the addition of FAD or FMN. The only effective substrate for the GLDHase was L-galactono-gamma-lactone.
authors.affiliation: Department of Food Biochemistry, Nagoya Women's University.
authors.forename: K
authors.initials: K
authors.lastname: Oba
authors.forename: S
authors.initials: S
authors.lastname: Ishikawa
authors.forename: M
authors.initials: M
authors.lastname: Nishikawa
authors.forename: H
authors.initials: H
authors.lastname: Mizuno
authors.forename: T
authors.initials: T
authors.lastname: Yamamoto
mesh_terms: D001205:Ascorbic Acid / Q000096:biosynthesis*; D004795:Enzyme Stability; D007700:Kinetics; D008928:Mitochondria / Q000201:enzymology*; D008970:Molecular Weight; D010088:Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification*; D044925:Oxidoreductases Acting on CH-CH Group Donors; D018517:Plant Roots / Q000201:enzymology*; D013379:Substrate Specificity; D014675:Vegetables / Q000201:enzymology*
chemical_list: D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; C093910:galactonolactone dehydrogenase; D001205:Ascorbic Acid