Purification and Characterization of Trehalase from Artemia Embryos and Larvae
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概要
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A soluble alkaline trehalase was purified from embryos and larvae of the brine shrimp, Artemia, by acetone treatment, chromatography on columns of DEAE-Sepharose Fast Flow, Con A-Sepharose and TSkgel AF-Chelate TOYOPEARL 650M, and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The purified enzyme subjected to SDS-PAGE showed a single protein band, suggesting a molecular mass of 70, 000 Da. The enzyme exhibited an apparent molecular mass of 58, 000 Da on gel filtration. Endoglycosidase H digestion of the enzyme did not affect the activity of the trehalase, and resulted in a molecular mass of 66, 000 Da on SDS-PAGE. The isoelectric point of the enzyme was estimated by gel electrofocusing to be approximately 4.7〜4.8. The catalytic activity showed a maximum at pH 8.0, and a specific activity of 140 μmoles glucose liberated from α, α-trehalose min^<-1> × mg^<-1> was observed at 30℃. The Km value for α, α-trehalose was estimated to be 8.4 mM. Among the eleven oligosaccharides and two α-glucoside derivatives studied, the enzyme hydrolyzed only α, α-trehalose. The enzyme was maximally active at 55℃ and had an activation energy of 55.8 kJ × mol^<-1>. The enzymatic reaction was completely inhibited by 0.1 mM HgCl_2. The activity of the purified enzyme was inhibited by 1 mM EDTA in the presence of 50 mM phosphate buffer, and the additions of appropriate amount of MnCl_2, MgCl_2 and CaCl_2 to the reaction mixture each protected the activity.
- 社団法人日本動物学会の論文
著者
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Tanaka Shin
Biology, Department of Medical Technology, School of Health Sciences, University of Occupational and
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Nambu Fumiko
Biology, Department of Medical Technology, School of Health Sciences, University of Occupational and
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Nambu Ziro
Biology, Department of Medical Technology, School of Health Sciences, University of Occupational and
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Nambu Fumiko
Biology, Department of Medical Technology, School of Health Sciences, University of Occupational and Environmental Heaalth
関連論文
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