title: Superoxide and hydrogen peroxide formation during enzymatic oxidation of DOPA by phenoloxidase.
abstract: Generation of superoxide anion and hydrogen peroxide during enzymatic oxidation of 3-(3,4-dihydroxyphenyl)-DL-alanine (DOPA) has been studied. The ability of DOPA to react with O2*- has been revealed. EPR spectrum of DOPA-semiquinone formed upon oxidation of DOPA by O2*- was observed using spin stabilization technique of ortho-semiquinones by Zn2+ ions. Simultaneously, the oxidation of DOPA by O2*- was found to produce hydrogen peroxide (H2O2). The analysis of H2O2 formation upon oxidation of DOPA by O2*- using 1-hydroxy-3-carboxy-pyrrolidine (CP-H), and SOD as competitive reagents for superoxide provides consistent values of the rate constant for the reaction between DOPA and O2*- being equal to (3.4+/-0.6)x10(5) M(-1) s(-1). The formation of H2O2 during enzymatic oxidation of DOPA by phenoloxidase (PO) has been shown. The H2O2 production was found to be SOD-sensitive. The inhibition of H2O2 production by SOD was about 25% indicating that H2O2 is produced both from superoxide anion and via two-electron reduction of oxygen at the enzyme. The attempts to detect superoxide production during enzymatic oxidation of DOPA using a number of spin traps failed apparently due to high value of the rate constant for DOPA interaction with O2*-.
authors.affiliation: Institute of Chemical Kinetics and Combustion, SB RAS, 630090, Novosibirsk, Russia. komarov@kinetics.nsc.ru
authors.forename: Denis A
authors.initials: DA
authors.lastname: Komarov
authors.forename: Irina A
authors.initials: IA
authors.lastname: Slepneva
authors.forename: Viktor V
authors.initials: VV
authors.lastname: Glupov
authors.forename: Valery V
authors.initials: VV
authors.lastname: Khramtsov
mesh_terms: D016227:Benzoquinones / Q000737:chemistry; D004295:Dihydroxyphenylalanine / Q000737:chemistry / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D014442:Monophenol Monooxygenase / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D019097:Spin Trapping; D013481:Superoxides / Q000737:chemistry / Q000378:metabolism*
chemical_list: D016227:Benzoquinones; D013481:Superoxides; C025232:semiquinone radicals; D004295:Dihydroxyphenylalanine; D006861:Hydrogen Peroxide; D014442:Monophenol Monooxygenase