Purification and Characterization of Wheat (Triticum aestivum) Polyphenol Oxidase
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概要
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Polyphenol oxidase was purified to homogeneity from Japanese wheat (Triticum aestivum cv. Tohoku 198) bran, using 4-methylcatechol as a substrate. The enzyme was purified 1670-fold with a recovery of 0.175% by alkalization, ammonium sulfate precipitation, anion exchange chromatography, hydrophobic chromatography and gel filtration chromatography. The molecular mass was estimated to be 35kDa and 40kDa by gel filtration chromatography and SDS-PAGE, respectively. The N-terminal amino acid sequence matched well with the internal sequences of other plant PPOs. The pH optima for the purified enzyme and the crude enzyme were 4.5 and 5.3, respectively. The purified enzyme was stable in the range of pH 6.5-7.5 at 4°C for 24hr. The purified enzyme was stable after heat treatment at 60°C for 10min. The purified enzyme oxidized L-tyrosine, pyrocatechol, 4-methylcatechol, L-DOPA, chlorogenic acid, (+)-catechin and (-)-epicatechin. The Km values of the purified enzyme for 4-methylcatechol and L-DOPA were 3.3mM and 8.3mM, respectively. The purified enzyme was strongly inhibited by dietyldithiocarbamate, thiourea, sodium azide, cystein, idoacetic acid, sodium fluoride, trans-cinnamic acid, p-coumaric acid, ferulic acid and vanillic acid. The KI value of the purified enzyme for ferulic acid was 108μM.
- 2005-02-01
著者
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KOMAE Kozo
Department of Wheat and Barley Research, National Institute of Crop Science
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Kihara Tomoko
Department of Food and Nutritional Sciences, College of Bioscience and Biotechnology, Chubu Universi
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KANEKO Shigenobu
Department of Wheat and Barley, National Institute of Crop Science, National Agricultural Research O
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MURATA Masatsune
Department of Nutrition and Food Science, Ochanomizu University
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Kihara Tomoko
Department Of Food And Nutritional Sciences College Of Bioscience And Biotechnology Chubu University
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Kihara Tomoko
Department Of Nutritional And Food Science Ochanomizu University
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HOMMA Seiichi
Department of Nutrition and Food Science, Ochanomizu University
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Kaneko Shigenobu
Department Of Agricultural Chemistry The University Of Tokyo
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Kaneko Shigenobu
Department Of Wheat And Barley Research National Institute Of Crop Science
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Homma Seiichi
Department Of Nutritional And Food Science Ochanomizu University
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Homma Seiichi
Department Of Nutrition & Food Science Ochanomizu University
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Komae Kozo
Department Of Food And Nutrition Faculty Of Science Of Living Osaka City University
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Murata Masatsune
Department Of Nutrition And Food Science Ochanomizu University
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Murata Masatsune
Department Of Nutritional And Food Science Ochanomizu University
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Murata Masatsune
Department Of Nutrition & Food Science Ochanomizu University
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HOMMA Seiichi
Department of Bioscience, Tokyo University of Agriculture
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