Cloning and characterization of a novel phytase from wastewater treatment yeast Hansenula fabianii J640 and expression in Pichia pastoris(ENVIRONMENTAL BIOTECHNOLOGY)
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概要
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Phosphohydrolysis of organic phosphorus compounds by acid phosphatases (EC3.1.3.1 and EC3.1.3.2) is an important method for efficient removal of phosphorus from high concentration organic wastewater. Another important method is supplementation of animal feed with phytase (EC3.1.3.8 and EC3.1.3.26), which improves the availability of phytate-phosphates (phosphate that are hydrolyzed by phytases), making it possible to add less phosphate to animal feed and resulting in the excretion of less phosphorus by the animals. In the present study, we purified a novel phytase from the wastewater treatment yeast Hansenula fabianii J640 (Hfphytase), cloned the 1456bp open reading frame (ORF) encoding Hfphytase, and characterized Hfphytase. The molecular weight of Hfphytase after deglycosylation by PNGaseF was 49kDa. The optimal pH and temperature for enzyme activity were 4.5 and 50℃, respectively. Hfphytase exhibits 40% identity with Debaryomyces castellii phytase, 37% identity with Aspergillus niger PhyB, and 34% identity with Saccharomyces cerevisiae Pho5p. Recombinant Hfphytase was transformed and expressed in Pichia pastoris. The yield was 23g/l by jar fermenter cultivation. The marked phosphohydrolysis activity exhibited by Hfphytase on six substrates (pNP-P, sodium phytate, glucose-1 phosphate, glucose-6 phosphate, α-glycerophosphate and β-glycerophosphate) indicated that it is a non-specific acid phosphatase.
著者
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Masaki Kazuo
National Research Institute of Brewing
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正木 和夫
酒総研
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正木 和夫
独立行政法人酒類総合研究所
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正木 和夫
北大・理・生物科学
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正木 和夫
(独)酒類総合研究所醸造技術応用研究部門
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正木 和夫
醸造技術応用研究部門
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Masaki Kazuo
National Res. Inst. Of Brewing 3-7-1 Kagamiyama Higashihiroshima Hiroshima 739-0046 Jpn
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Fujii Tsutomu
National Research Institute Of Brewing:graduate School Of Biosphere Sciences Hiroshima University
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WATANABE Takashi
Graduate School of Biomedical Engineering, Tohoku University
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Ikeda Hiroko
National Research Institute of Brewing
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Fujii Tsutomu
Graduate School of Biosphere Science, Hiroshima University
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Iefuji Haruyuki
Graduate School of Biosphere Science, Hiroshima University
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Iefuji Haruyuki
National Res. Inst. Of Brewing
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Iefuji Haruyuki
National Research Institute Of Brewing:graduate School Of Biosphere Sciences Hiroshima University
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Iefuji Haruyuki
Graduate School Of Biosphere Sci. Hiroshima Univ.
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Ikeda Hiroko
National Epilepsy Center Shizuoka
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Watanabe Takashi
Graduate School Of Biosphere Science Hiroshima University:national Research Institute Of Brewing
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Watanabe Takashi
Graduate Department Of Mathematical & Life Sciences Hiroshima Univ.
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正木 和夫
広島大院・生物圏:酒総研
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