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Efficient Production of 2-O-α-D-Glucosyl Glycerol Catalyzed by an Engineered Sucrose Phosphorylase from Bifidobacterium longum

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Abstract

2-O-α-D-Glucosyl glycerol (2-αGG) can be used as a multipurpose anti-aging, cell-stimulating, and skin moisturizing agent in the cosmetic industry. Sucrose phosphorylase (SPase) has been widely used in the production of 2-αGG. In this paper, the gene encoding sucrose phosphorylase from Bifidobacterium longum (BlSP) was inserted into pRSF-Duet-1 to construct the recombinant plasmid pRSF-BlSP and was functionally expressed in E. coli BL21(DE3) to be used as a biocatalyst for the synthesis of 2-αGG firstly. The mutations of BlSP were carried out based on alanine scanning, and a positive mutant G293A with a 50% increase in activity for 2-αGG production was identified. Mutant G293A has less Km and bigger kcat/Km towards glycerol than the parental BlSP. Subsequently, the production of 177.6 g/L 2-αGG was attained from 1 M sucrose and 1.2 M glycerol catalyzed by 17 mg/mL G293A mutant. This study indicated that BlSP has good potential in the production of 2-αGG.

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Data Availability

All proteins in the present study are available from UniProtKB via the accession codes. The original data are provided with the paper.

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Funding

This work was financed by the National Key R & D Program of China (2021YFC2101500), NSFC (21878155), the Jiangsu Synergetic Innovation Center, or Advanced Bio-manufacture, and PAPD.

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Authors and Affiliations

  1. College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, 211816, China

    Jiping Lei, Kexin Tang, Ting Zhang, Yan Li, Zhen Gao & Honghua Jia

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  1. Jiping Lei
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  2. Kexin Tang
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  3. Ting Zhang
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  4. Yan Li
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  5. Zhen Gao
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  6. Honghua Jia
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Contributions

J.P.L. conducted most of the experiments. Y.L. and H.H.J. designed and supervised the project. Z.G. provided experimental reagents and equipment. K.X.T. conducted kinetic simulation analysis of enzymes. T.Z. performed activity assay and tested the related reactivity of substrates. All the authors discussed the design and results, commented on the manuscript, and approved the manuscript.

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Correspondence to Yan Li or Honghua Jia.

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Lei, J., Tang, K., Zhang, T. et al. Efficient Production of 2-O-α-D-Glucosyl Glycerol Catalyzed by an Engineered Sucrose Phosphorylase from Bifidobacterium longum. Appl Biochem Biotechnol 194, 5274–5291 (2022). https://doi.org/10.1007/s12010-022-03939-z

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