Biocatalytic Synthesis of Human Milk Oligosaccharides Lacto-N-Biose Via Modular Enzymatic Cascade with ATP Regeneration | AIChE

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Biocatalytic Synthesis of Human Milk Oligosaccharides Lacto-N-Biose Via Modular Enzymatic Cascade with ATP Regeneration

Authors 

Du, Z. - Presenter, Shandong University
Liu, Z., Shandong University
Tan, Y., Shandong University
Fang, X., Shandong University
Human milk oligosaccharides (HMOs), the third most abundant solid component of human milk, are reported to be beneficial to infant health (Eriksen et al., 2018; Han et al., 2012). The biosynthesis of Lacto-N-biose (LNB), the building block for HMOs, suffers from excessive cofactors addition and intermediate inhibition (Nishimoto and Kitaoka, 2007; Nishimoto 2020). Here, we developed an in vitro multienzyme cascade composed of LNB module, ATP regeneration, and pyruvate oxidase-driven phosphate recycling to produce LNB. The integration between ATP regeneration and Pi alleviation increased the LNB conversion ratio and resulted in a ΔG'° decrease of 540 KJ/mol. Under the optimal conditions, the LNB conversion ratio was improved from 0.34 to 0.83 mol/mol GlcNAc and the ATP addition decreased to 50%. Finally, about 0.96 mol/mol GlcNAc and 71.6 mg LNB g-1 GlcNAc h-1 of LNB yield were achieved in 100 mL reaction system. The synergistic strategy not only paves the way for producing LNB but also facilitates other chemicals with multienzyme cascades.

Reference:

Du, Z. Q.; Liu, Z. Y.; Tan, Y. S.; Niu, K. L.; Guo, W.; Jia, Y.Y.; Fang, X. (2021). Lacto-N-biose synthesis via a modular enzymatic cascade with ATP regeneration. iScience, 24(3), 102236.

Eriksen, K. G.; Christensen, S. H.; Lind, M. V.; and Michaelsen, K. F. (2018). Human milk composition and infant growth. Curr. Opin. Clin. Nutr. 21 (3), 200-206.

Han, N. S.; Kim, T. J.; Park, Y. C.; Kim, J.; and Seo, J. H. (2012). Biotechnological production of human milk oligosaccharides. Biotechnol. Adv. 30 (6), 1268-1278.

Nishimoto, M. (2020). Large scale production of lacto-N-biose I, a building block of type I human milk oligosaccharides, using sugar phosphorylases. Biosci. Biotech. Bioch. 84, 17-24.

Nishimoto, M. and Kitaoka, M. (2007). Practical preparation of lacto-N-biose I, a candidate for the Bifidus factor in human milk. Biosci. Biotech. Bioch. 71 (8), 2101-2104.

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