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Metabolic Engineering of De Novo Pathway for the Production of 2′-Fucosyllactose in Escherichia coli

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Abstract

2′-Fucosyllactose (2′-FL), one of the most abundant oligosaccharides in human milk, has gained increased attention owing to its nutraceutical and pharmaceutical potential. However, limited availability and high-cost of preparation have limited its widespread application and in-depth investigation of its potential functions. Here, a modular pathway engineering was implemented to construct an Escherichia coli strain to improve the biosynthesis titer of 2′-FL. Before overexpression of manB, manC, gmd, wcaG, and heterologous expression of futC, genes wcaJ and lacZ encoding UDP-glucose lipid carrier transferase and β-galactosidase, respectively, were inactivated from E. coli BL21 (DE3) with the CRISPR-Cas9 system, which inhibited the production of 2′-FL. The results showed that final shake flask culture yielded a 3.8-fold increase in 2′-FL (0.98 g/L) from the engineered strain ELC07. Fed-batch fermentation conditions were optimized in a 3-L bioreactor. The highest titer of 2′-FL (18.22 g/L) was obtained, corresponding to a yield of 0.25 g/g glycerol and a substrate conversion of 0.88 g/g lactose.

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Data Availability

All data generated or analyzed during this study are included in this published article and its supplementary information files.

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Funding

This work was financially supported by Major Science and Technology Innovation Project of Shandong Province (Grant No. 2020CXGC010601) and Kitty Hawk Project of Zhejiang Provincial Administration for Market Regulation (Grant No. CY2022004).

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Authors and Affiliations

  1. State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214122, Jiangsu, China

    Chenchen Li, Mengli Li, Miaomiao Hu & Tao Zhang

  2. International Joint Laboratory on Food Safety, Jiangnan University, Wuxi, 214122, Jiangsu, China

    Tao Zhang

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Contributions

All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by CL and ML. The first draft of the manuscript was written by CL. Reviewing and editing of the manuscript were performed by ML and MH. Supervision, giving major comments, and project administration were contributed by TZ. All authors read and approved the final manuscript.

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Correspondence to Tao Zhang.

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Li, C., Li, M., Hu, M. et al. Metabolic Engineering of De Novo Pathway for the Production of 2′-Fucosyllactose in Escherichia coli. Mol Biotechnol 65, 1485–1497 (2023). https://doi.org/10.1007/s12033-023-00657-7

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