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Superoxide anion is the initial product in the hydrogen peroxide formation catalyzed by NADPH oxidase in porcine thyroid plasma membrane.

Abstract

The plasma membrane fraction from porcine thyroid is known to exhibit an NADPH-dependent production of hydrogen peroxide (H2O2), which is utilized for the oxidative biosynthesis of thyroid hormones catalyzed by thyroid peroxidase. The H2O2 formation is cyanide-insensitive, ATP-activatable, and Ca2+-dependent (Nakamura, Y., Ogihara, S., and Ohtaki, S. (1987) J. Biochem. (Tokyo) 102, 1121-1132). It remains unknown, however, whether H2O2 is produced directly from molecular oxygen (O2) or formed via dismutation of superoxide anion (O2-). We therefore attempted to analyze the mechanism of H2O2 formation by utilizing a new method for the simultaneous measurement of O2- and H2O2, in which diacetyldeuteroheme-substituted horseradish peroxidase was employed as the trapping agent for both oxygen metabolites. When NADPH was incubated with the membrane fraction in the presence of the heme-substituted peroxidase, a massive O2 consumption was observed together with the formation of compound III, and O2- adduct of the peroxidase. The amounts of compound III formed and O2 consumed were stoichiometric with each other, while formation of compound II, an indicative of H2O2, was not observed during the reaction. On the other hand, when an excess amount of superoxide dismutase was included in the reaction mixture, compound II was produced with complete suppression of the compound III formation. NADH minimally supported both O2 consumption and formation of compound III or II. These results indicate that the NADPH oxidase in the plasma membrane of thyroid produces O2- as the primary metabolite of O2 and hence that H2O2 required for the thyroid hormone synthesis provided through the dismutation of O2-.

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  • Authors+Show Affiliations

    ,

    Central Laboratory for Clinical Investigation, Miyazaki Medical College Hospital, Japan.

    , , ,

    Source

    The Journal of biological chemistry 264:9 1989 Mar 25 pg 4759-61

    MeSH

    Animals
    Catalysis
    Cell Membrane
    Diacetyl
    Hemeproteins
    Hydrogen Peroxide
    NADH, NADPH Oxidoreductases
    NADPH Oxidases
    Oxygen Consumption
    Spectrophotometry
    Superoxides
    Swine
    Thyroid Gland

    Pub Type(s)

    Journal Article
    Research Support, Non-U.S. Gov't

    Language

    eng

    PubMed ID

    2538459

    Citation

    Nakamura, Y, et al. "Superoxide Anion Is the Initial Product in the Hydrogen Peroxide Formation Catalyzed By NADPH Oxidase in Porcine Thyroid Plasma Membrane." The Journal of Biological Chemistry, vol. 264, no. 9, 1989, pp. 4759-61.
    Nakamura Y, Ohtaki S, Makino R, et al. Superoxide anion is the initial product in the hydrogen peroxide formation catalyzed by NADPH oxidase in porcine thyroid plasma membrane. J Biol Chem. 1989;264(9):4759-61.
    Nakamura, Y., Ohtaki, S., Makino, R., Tanaka, T., & Ishimura, Y. (1989). Superoxide anion is the initial product in the hydrogen peroxide formation catalyzed by NADPH oxidase in porcine thyroid plasma membrane. The Journal of Biological Chemistry, 264(9), pp. 4759-61.
    Nakamura Y, et al. Superoxide Anion Is the Initial Product in the Hydrogen Peroxide Formation Catalyzed By NADPH Oxidase in Porcine Thyroid Plasma Membrane. J Biol Chem. 1989 Mar 25;264(9):4759-61. PubMed PMID: 2538459.
    * Article titles in AMA citation format should be in sentence-case
    TY - JOUR T1 - Superoxide anion is the initial product in the hydrogen peroxide formation catalyzed by NADPH oxidase in porcine thyroid plasma membrane. AU - Nakamura,Y, AU - Ohtaki,S, AU - Makino,R, AU - Tanaka,T, AU - Ishimura,Y, PY - 1989/3/25/pubmed PY - 1989/3/25/medline PY - 1989/3/25/entrez SP - 4759 EP - 61 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 264 IS - 9 N2 - The plasma membrane fraction from porcine thyroid is known to exhibit an NADPH-dependent production of hydrogen peroxide (H2O2), which is utilized for the oxidative biosynthesis of thyroid hormones catalyzed by thyroid peroxidase. The H2O2 formation is cyanide-insensitive, ATP-activatable, and Ca2+-dependent (Nakamura, Y., Ogihara, S., and Ohtaki, S. (1987) J. Biochem. (Tokyo) 102, 1121-1132). It remains unknown, however, whether H2O2 is produced directly from molecular oxygen (O2) or formed via dismutation of superoxide anion (O2-). We therefore attempted to analyze the mechanism of H2O2 formation by utilizing a new method for the simultaneous measurement of O2- and H2O2, in which diacetyldeuteroheme-substituted horseradish peroxidase was employed as the trapping agent for both oxygen metabolites. When NADPH was incubated with the membrane fraction in the presence of the heme-substituted peroxidase, a massive O2 consumption was observed together with the formation of compound III, and O2- adduct of the peroxidase. The amounts of compound III formed and O2 consumed were stoichiometric with each other, while formation of compound II, an indicative of H2O2, was not observed during the reaction. On the other hand, when an excess amount of superoxide dismutase was included in the reaction mixture, compound II was produced with complete suppression of the compound III formation. NADH minimally supported both O2 consumption and formation of compound III or II. These results indicate that the NADPH oxidase in the plasma membrane of thyroid produces O2- as the primary metabolite of O2 and hence that H2O2 required for the thyroid hormone synthesis provided through the dismutation of O2-. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/2538459/Superoxide_anion_is_the_initial_product_in_the_hydrogen_peroxide_formation_catalyzed_by_NADPH_oxidase_in_porcine_thyroid_plasma_membrane_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=2538459 DB - PRIME DP - Unbound Medicine ER -