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A Novel Glucose-Tolerant GH1 β-Glucosidase and Improvement of Its Glucose Tolerance Using Site-Directed Mutation

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Abstract

A novel GH1 β-glucosidase gene (bgla) from marine bacterium was sequenced and expressed in Escherichia coli. After purification by Ni2+ affinity chromatography, the recombinant protein was characterized. The purified recombinant enzyme showed maximum activity at 40 °C, pH 7.5 and was stable between temperatures that range from 4 to 30 °C and over the pH range of 6–10. The enzyme displayed a high tolerance to glucose and maximum stimulation at the presence of 100 mM glucose. To improve glucose tolerance of the enzyme, a site-directed mutation (f171w) was introduced into β-glucosidase. The recombinant F171W showed a higher glucose tolerance than the wild type and maintained more than 40% residual activity at the presence of 4 M glucose. Additionally, the recombinant enzymes showed notable tolerance to ethanol. These properties suggest the enzymes may have potential applications for the fermentation of lignocellulosic sugars and the production of biofuels.

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Funding

This study was funded by the National Natural Science Foundation of China (31900035); the Central Public-interest Scientific Institution Basal Research Fund, YSFRI, CAFS (No. 20603022018006); the Central Public-interest Scientific Institution Basal Research Fund, CAFS (NO.2020TD67); the Financial Fund of the Ministry of Agriculture, P. R. China (No.NFZX2013); and the Scientific and the Technological Innovation Project Financially Supported by Qingdao National Laboratory for Marine Science and Technology (No. 2016ASKJ14).

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Authors and Affiliations

  1. Key Laboratory of Sustainable Development of Polar Fishery, Ministry of Agriculture and Rural Affairs, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao, 266071, China

    Jingjing Sun, Wei Wang & Jianhua Hao

  2. Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266071, China

    Jingjing Sun, Wei Wang & Jianhua Hao

  3. Qingdao Institute for Food and Drug Control, Qingdao, 266071, China

    Yu Ying

  4. Jiangsu Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resource, Lianyungang, 222005, China

    Jianhua Hao

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  1. Jingjing Sun
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  2. Wei Wang
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  3. Yu Ying
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  4. Jianhua Hao
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Contributions

J. Sun and J. Hao contributed to the conception and design of the study; J. Sun contributed to the Data curation; J. Sun, J. Hao, and W. Wang contributed significantly to analysis and manuscript preparation; J. Sun and Y. Ying performed writing-review and editing.

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Correspondence to Jingjing Sun or Jianhua Hao.

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Sun, J., Wang, W., Ying, Y. et al. A Novel Glucose-Tolerant GH1 β-Glucosidase and Improvement of Its Glucose Tolerance Using Site-Directed Mutation. Appl Biochem Biotechnol 192, 999–1015 (2020). https://doi.org/10.1007/s12010-020-03373-z

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