| Title |
Characterization of a Novel β-Glucosidase-Like Activity from a Soil Metagenome |
| Author |
Chengjian Jiang1, Gefei Ma1, Shuangxi Li1, Tingting Hu1, Zhiqun Che1, Peihong Shen1, Bing Yan2, and Bo Wu1* |
| Address |
1College of Life Science and Technology, Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, The Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, Guangxi University, Guangxi 530004, P. R. China, 2Guangxi Key Laboratory for Marine Biotechnology, Guangxi Institute of Oceanology, Guangxi 536000, P. R. China |
| Bibliography |
Journal of Microbiology, 47(5),542-548, 2009,
|
| DOI |
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| Key Words |
uncultured soil microorganisms, function-based screening strategy, β-glucosidase |
| Abstract |
We report the cloning of a novel β-glucosidase-like gene by function-based screening of a metagenomic library from uncultured soil microorganisms. The gene was named bgl1C and has an open reading frame of 1,443 base pairs. It encodes a 481 amino acid polypeptide with a predicted molecular mass of about 57.8 kDa. The deduced amino acid sequence did not show any homology with known β-glucosidases. The putative β-glucosidase gene was subcloned into the pETBlue-2 vector and overexpressed in E. coli Tuner (DE3) pLacІ; the recombinant protein was purified to homogeneity. Functional characterization with a high performance
liquid chromatography method demonstrated that the recombinant Bgl1C protein hydrolyzed D-glucosyl-β-(1-4)-D-glucose to glucose. The maximum activity for Bgl1C protein occurred at pH 8.0 and 42°C using p-nitrophenyl-β-D-glucoside as the substrate. A CaCl2 concentration of 1 mM was required for optimal activity. The putative β-glucosidase had an apparent Km value of 0.19 mM, a Vmax value of 4.75
U/mg and a kcat value of 316.7/min under the optimal reaction conditions. The biochemical characterization of Bgl1C has enlarged our understanding of the novel enzymes that can be isolated from the soil metagenome. |
