Characterization of a recombinant amylolytic enzyme of hyperthermophilic archaeon Thermofilum pendens with extremely thermostable maltogenic amylase activity.

Xiaolei Li, Dan Li, Yongguang Yin, Kwan-Hwa Park

Key Laboratory of Agricultural Products Processing in Jilin Province Universities, Changchun University, Changchun 130022, People's Republic of China.

Applied microbiology and biotechnology 2010 Feb

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A gene (Tpen_1458) encoding a putative alpha amylase from hyperthermophilic archaeon Thermofilum pendens (TfMA) was cloned and expressed in Escherichia coli. The recombinant amylolytic enzyme was purified by Ni-NTA affinity chromatography and its catalytic properties were examined. Purified TfMA was extremely thermostable with a half-life of 60 min at an optimal temperature of 95 degrees C. TfMA activity increased to 136% in the presence of 5 mM CaCl(2). Maximal activity was measured toward gamma-cyclodextrin with a specific activity of 56 U/mg using copper bicinchoninate method. TfMA catalyzed the ring-opening reaction by cleaving one alpha-1,4-glycosidic linkage of cyclodextrin to produce corresponding single maltooligosaccharide at the initial time. The final products from cyclodextrins, linear maltooligosaccharides, and starch were glucose and maltose, and TfMA could also degrade pullulan and amylase inhibitor acarbose to panose and acarviosine-glucose, respectively. These results revealed that TfMA is a novel maltogenic amylase.

Citation

Xiaolei Li, Dan Li, Yongguang Yin, Kwan-Hwa Park. Characterization of a recombinant amylolytic enzyme of hyperthermophilic archaeon Thermofilum pendens with extremely thermostable maltogenic amylase activity. Applied microbiology and biotechnology. 2010 Feb;85(6):1821-30