A mechanistic study of the formation of hydroxyl radicals induced by horseradish peroxidase with NADH.
During the oxidation of NADH by horseradish peroxidase (HRP-Fe(3+)), superoxide (O(-)(2)) is produced, and HRP-Fe(3+) is converted to compound III. Superoxide dismutase inhibited both the generation of O(-)(2) and the formation of compound III. In contrast, catalase inhibited only the generation of O(-)(2). Under anaerobic conditions, the formation of compound III did not occur in the presence of NADH, thus indicating that compound III is produced via formation of a ternary complex consisting of HRP-Fe(3+), NADH and oxygen. The generation of hydroxyl radicals was dependent upon O(-)(2) and H(2)O(2) produced by HRP-Fe(3+)-NADH. The reaction of compound III with H(2)O(2) caused the formation of compound II without generation of hydroxyl radicals. Only HRP-Fe(3+)-NADH (but not K(+)O(-)(2) and xanthine oxidase-hypoxanthine) was able to induce the conversion of metmyoglobin to oxymyoglobin, thus suggesting the participation of a ternary complex made up of HRP-Fe(2+…)O(2)(…)NAD(.) (but not free O(-)(2) or H(2)O(2)) in the conversion of metmyoglobin to oxymyoglobin. It appears that a cyclic pathway is formed between HRP-Fe(3+), compound III and compound II in the presence of NADH under aerobic conditions, and a ternary complex plays the central roles in the generation of O(-)(2) and hydroxyl radicals.
Department of Biology in Hokkaido Pharmaceutical University School of Pharmacy, Katsuraoka-cho7-1, Otaru, 0470264 Japan. miurat@hokuyakudai.ac.jp
Toshiaki
T
Miura
D002374:Catalase / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D006735:Horseradish Peroxidase / Q000737:chemistry* / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000737:chemistry; D017665:Hydroxyl Radical / Q000737:chemistry* / Q000378:metabolism; D008786:Metmyoglobin / Q000378:metabolism; D009211:Myoglobin / Q000378:metabolism; D009243:NAD / Q000737:chemistry* / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism
D009211:Myoglobin; C017595:oxymyoglobin; D009243:NAD; D013481:Superoxides; D008786:Metmyoglobin; D017665:Hydroxyl Radical; D006861:Hydrogen Peroxide; D006735:Horseradish Peroxidase; D002374:Catalase; D013482:Superoxide Dismutase