Volume 15, Issue 9 e202102539
Research Article

Substrate and Process Engineering for Biocatalytic Synthesis and Facile Purification of Human Milk Oligosaccharides

Yuanyuan Bai

Yuanyuan Bai

Department of Chemistry, University of California, Davis, One Shields Avenue, 95616 Davis, California, USA

Search for more papers by this author
Dr. Xiaohong Yang

Dr. Xiaohong Yang

Department of Chemistry, University of California, Davis, One Shields Avenue, 95616 Davis, California, USA

Search for more papers by this author
Dr. Hai Yu

Dr. Hai Yu

Department of Chemistry, University of California, Davis, One Shields Avenue, 95616 Davis, California, USA

Search for more papers by this author
Prof. Dr. Xi Chen

Corresponding Author

Prof. Dr. Xi Chen

Department of Chemistry, University of California, Davis, One Shields Avenue, 95616 Davis, California, USA

Search for more papers by this author
First published: 31 January 2022
Citations: 5

Graphical Abstract

Got milk sugars? A highly efficient multistep one-pot multienzyme (MSOPME) strategy is developed for carbohydrate synthesis by integrating biocatalysis substrate and process engineering concepts. A single C18-cartridge purification process is sufficient for any target. The strategy is demonstrated for preparative and gram-scale synthesis of numerous human milk oligosaccharides (HMOs), which are attractive targets for industrial biocatalytic production.

Abstract

Innovation in process development is essential for applying biocatalysis in industrial and laboratory production of organic compounds, including beneficial carbohydrates such as human milk oligosaccharides (HMOs). HMOs have attracted increasing attention for their potential application as key ingredients in products that can improve human health. To efficiently access HMOs through biocatalysis, a combined substrate and process engineering strategy is developed, namely multistep one-pot multienzyme (MSOPME) design. The strategy allows access to a pure tagged HMO in a single reactor with a single C18-cartridge purification process, despite the length of the target. Its efficiency is demonstrated in the high-yielding (71–91 %) one-pot synthesis of twenty tagged HMOs (83–155 mg), including long-chain oligosaccharides with or without fucosylation or sialylation up to nonaoses from a lactoside without the isolation of the intermediate oligosaccharides. Gram-scale synthesis of an important HMO derivative – tagged lacto-N-fucopentaose-I (LNFP-I) – proceeds in 84 % yield. Tag removal is carried out in high efficiency (94–97 %) without the need for column purification to produce the desired natural HMOs with a free reducing end. The method can be readily adapted for large-scale synthesis and automation to allow quick access to HMOs, other glycans, and glycoconjugates.

Conflict of interest

The authors declare no conflict of interest.

Data Availability Statement

The data that support the findings of this study are available in the supplementary material of this article.