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Enhancing thermostability and activity of sucrose phosphorylase for high-level production of 2-O-α-d-glucosylglycerol

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Abstract

Sucrose phosphorylase (SPase) can transfer the glucosyl group of sucrose to different compounds and has been widely used in industry. To overcome the low thermostability of the sucrose phosphorylase from Leuconostoc mesenteroides ATCC 12291 (LmSP), a method named PROSS was used to construct mutants with increased thermostability. All variants were screened by measuring their residual activities after heating at 50°C. Then, a single point mutant and a combined mutant with improved thermostability and activity were obtained. The half-lives of mutants at 50°C were approximately twice as high as those of the wild type. In addition, 2-O-α-d-glucosylglycerol (αGG) was synthesized by the wild type and the two improved variants, and the reaction conditions were optimized. Under the conditions of glycerol concentration of 3.2 mol/L, sucrose concentration of 1.2 mol/L, and enzyme concentration of 40 U/mL at 37°C for 60 h, the yield of αGG reached the maximum, and the sucrose conversion rate of the wild type, the mutant V23L and the combined mutant V23L/S424R were 62.3%, 70.7% and 76.3%, respectively. In this study, SPase mutants with higher activity and stability were obtained, and achieved high-level production of αGG.

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Acknowledgements

This study was funded by the Key Research and Development Program of China (2021YFC2100102-03), the National Natural Science Foundation of China (32001064). The computational results used in this article were obtained using Interdisciplinary Center for Modern Technologies facilities, NCU, Torun, Poland.

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Authors and Affiliations

  1. Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, China

    Linli Yang, Yujuan Shen, Haiquan Yang, Xianzhong Chen, Wei Shen & Yuanyuan Xia

  2. Institute of Physics, Faculty of Physics, Astronomy and Informatics, Nicolaus Copernicus University in Torun, Grudziadzka 5, 87-100, Torun, Poland

    Lukasz Peplowski

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  1. Linli Yang
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Contributions

LLY and YYX: designed the research. LLY, YYX, and YJS: performed the research and analyzed the data. PL: ran the MD simulations and analyzed the data, YYL: wrote the paper. HQY, WS, XZC and YYX: supervised the research work.

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Correspondence to Wei Shen or Yuanyuan Xia.

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Yang, L., Peplowski, L., Shen, Y. et al. Enhancing thermostability and activity of sucrose phosphorylase for high-level production of 2-O-α-d-glucosylglycerol. Syst Microbiol and Biomanuf 2, 643–652 (2022). https://doi.org/10.1007/s43393-022-00090-y

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  • DOI: https://doi.org/10.1007/s43393-022-00090-y

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