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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/27027

Title: Cloning, Expression, and Characterization of an Enzyme Possessing both Glutaredoxin and Dehydroascorbate Reductase Activity from Taiwanofungus camphorata
Authors: Chuian-Fu Ken;Choa-Yi Lin;Yu-Chi Jiang;Lisa Wen;Chi-Tsai Lin
Contributors: NTOU:Institute of Bioscience and Biotechnology
國立臺灣海洋大學:生物科技研究所
Keywords: Taiwanofungus camphorata;glutaredoxin;three-dimension homology structure (3D homology structure);expression;β-hydroxyethyl disulfide [HED, (HOCH2CH2)2S2];dehydroascorbate (DHA)
Date: 2009
Issue Date: 2011-10-21T02:22:21Z
Publisher: Journal of Agricultural and Food Chemistry
Abstract: Abstract:Glutaredoxins (Grxs) play important roles in the reduction of disulfides via reduced glutathione as a reductant. A cDNA (503 bp, EU193660) encoding a putative Grx was cloned from Taiwanofugus camphorata (Tc). The deduced amino acid sequence is conserved among the reported dithiol Grxs. A 3D homology structure was created for this TcGrx. To characterize the TcGrx enzyme, the coding region was subcloned into an expression vector pET-20b(+) and transformed into Escherichia coli. Functional TcGrx was expressed and purified by Ni2+-nitrilotriacetic acid Sepharose. The purified enzyme showed bands of 15 kDa on 15% sodium dodecyl sulfate−polyacrylamide gel electrophoresis (SDS−PAGE). The TcGrx encodes a protein possessing both Grx and dehydroascorbate reductase (DHAR) activity. The Michaelis constant (Km) values for β-hydroxyethyl disulfide (HED) and dehydroascorbate (DHA) were 0.57 and 1.85 mM, respectively. The half-life of deactivation of the protein at 100 °C was 8.5 min, and its thermal inactivation rate constant Kd was 6.52 × 10−2 min−1. The enzyme was active under a broad pH range from 6.0 to 10.0 and in the presence of imidazole up to 0.4 M. The enzyme was susceptible to SDS denaturation and protease degradation/inactivation.
Relation: 57(21), pp.10357–10362
URI: http://ntour.ntou.edu.tw/handle/987654321/27027
Appears in Collections:[生命科學暨生物科技學系] 期刊論文

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