Characterization and application of an acidophilic and thermostable β-glucosidase from Thermofilum pendens(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)
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概要
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The gene encoding a β-glucosidase from the archaeon Thermoflium pendens (Tpbgl) was cloned and expressed in Escherichia coli. The purified recombinant enzyme had a molecular mass of 77.8 kDa and released glucose or mannose from p-nitrophenyl-β-D-glucopyranoside (pNPG), cellobiose, mannobiose, and genistin. Peak Tpbgl activity was detected at 90℃, and 50% activity remained after incubation for 60 min at 95℃. The optimal pH for pNPG hydrolysis was 3.5. When the enzyme was incubated with pNPG in the presence of ethanol and propanol, the glucose moiety was transferred to acceptor alcohols. Tpbgl is the archaeal β-glucosidase from glucoside hydrolase family 3 and found to be most heat stable under extremely acidic conditions (pH 3.5). The kinetic parameters revealed that Tpbgl had the highest catalytic efficiency toward pNPG (k_<cat>/K_m = 3.05) with strong substrate affinity for such natural substrates as cellobiose (K_m = 0.149) and mannobiose (K_m = 0.147). Genistin solubilized in 10-40% DMSO was hydrolyzed to genistein with nearly 99% conversion, indicating that high concentrations of the water-insoluble isofiavone glycoside can be treated by the enzyme. Our results indicate that Tpbgl has great potential in cellulose saccharification and the glucoside hydrolysis of natural compounds.
- 公益社団法人日本生物工学会の論文
- 2013-05-00
著者
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Park Kwan-hwa
Department Of Food Science And Technology College Of Agriculture And Life Sciences Seoulnational Uni
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Park Sung-hoon
Department Of Chemical Engineering Pusan National University
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Park Kwan-Hwa
Department of Foodservice Management and Nutrition, Sangmyung University
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Li Dan
Key Laboratory of Agricultural Products Processing in Jilin Provincial Universities, Changchun University
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Li Xiaolei
Key Laboratory of Agricultural Products Processing in Jilin Provincial Universities, Changchun University
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Dang Wei
Key Laboratory of Agricultural Products Processing in Jilin Provincial Universities, Changchun University
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Tran Phuong
Department of Foodservice Management and Nutrition, Sangmyung University
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Oh Byung-Chul
Lee Gil Ya Cancer and Diabetes Institute, Gachon University of Medicine and Science
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Hong Wan-Soo
Department of Foodservice Management and Nutrition, Sangmyung University
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Lee Jin-Sil
Department of Foodservice Management and Nutrition, Sangmyung University
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Park Sung-Hoon
Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University
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- Characterization and application of an acidophilic and thermostable β-glucosidase from Thermofilum pendens(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)