Gene expression and molecular characterization of a thermostable trehalose phosphorylase fromThermoanaerobacter tengcongensis |
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Authors: | REN Yuanyuan Dai Xiuyu ZHOU Jian PEI Huadong Xiang Hua |
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Institution: | 1. State Key Laboratory of Microbial Resources and Center for Molecular Microbiology, Institute of Microbiology, Chinese Academy of Sciences, Beijing,100080, China;Graduate School of Chinese Academy of Sciences, Beijing,100037, China 2. State Key Laboratory of Microbial Resources and Center for Molecular Microbiology, Institute of Microbiology, Chinese Academy of Sciences, Beijing,100080, China |
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Abstract: | A gene encoding the trehalose phosphorylase (TreP), which reversibly catalyzes trehalose degradation and synthesis from α-glucose-1-phosphate
(α-Glc-1-P) and glucose, was cloned fromThermoanaerobacter tengcongensis and successfully expressed inEscherichia coli. The overexpressed TreP, with a molecular mass of approximately 90 kDa, was determined by SDS-PAGE. It catalyzes trehalose
synthesis and degradation optimally at 70°C (for 30 min), with the optimum pHs at 6.0 and 7.0, respectively. It is highly
thermostable, with a 77% residual activity after incubation at 50°C for 7 h. Under the optimum reaction conditions, 50 μg
crude enzyme of the TreP is able to catalyze the synthesis of trehalose up to 11.6 mmol/L from 25 mmol/L α-Glc-1-P and 125
mmol/L glucose within 30 min, while only 1.5 mmol/L out of 250 mmol/L trehalose is degraded within the same time period. Dot
blotting revealed that thetreP gene inT. tengcongensis was upregulated in response to salt stress but downregulated when trehalose was supplied. Both results indicate that the
dominant function of theT. tengcongensis TreP is catalyzing trehalose synthesis but not degradation. Thus it might provide a novel route for industrial production
of trehalose. |
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Keywords: | gene expression Thermoanaerobacter tengcongensis trehalose phosphorylase dot blot |
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