title: Oxymetric and spectrophotometric study of the ascorbate oxidase activity shown by frog epidermis tyrosinase.
abstract: Many studies concerning the effect of ascorbic acid on the action of tyrosinase on several substrates have been carried out with contradictory results. The results shown in this work comprise a hypothetical reaction mechanism, which explains the ascorbate oxidase activity of frog epidermis tyrosinase. The reaction between frog epidermis tyrosinase and L-ascorbic acid was studied by oxymetric and spectrophotometric assays. The activity was linearly related to enzyme concentration, with a Michaelis constant for L-ascorbic acid of 0.160 +/- 0.009 mM and Vmax of 90 +/- 4 nM/s. Maximum activity was obtained at pH 7.5. The stoichiometry of the reaction was calculated by measuring the substrate (O2 and L-ascorbic acid) consumption as well as the initial rates of the consumption of oxygen and the disappearance of L-ascorbic acid. The stoichiometry was found to be 1:2 (O2:L-ascorbic acid). The action of the tyrosinase inhibitor tropolone was also studied. All the results present evidence concerning the ascorbate oxidase activity of frog epidermis tyrosinase and a possible reaction mechanism based on the different enzymatic forms of tyrosinase to explain such activity.
authors.affiliation: Departamento de Bioquímica y Biología Molecular-A, Facultad de Biología, Universidad de Murcia, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Ramón Ros
authors.forename: J N
authors.initials: JN
authors.lastname: Rodríguez-López
authors.forename: J
authors.initials: J
authors.lastname: Carlos Espín
authors.forename: R
authors.initials: R
authors.lastname: Varón
authors.forename: F
authors.initials: F
authors.lastname: García-Cánovas
mesh_terms: D000818:Animals; D001001:Anura; D001204:Ascorbate Oxidase / Q000378:metabolism*; D001205:Ascorbic Acid / Q000378:metabolism*; D004789:Enzyme Activation; D004817:Epidermis / Q000201:enzymology*; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D001204:Ascorbate Oxidase; D014442:Monophenol Monooxygenase; D001205:Ascorbic Acid