Production of HMO by Chemoenzymatic Methods

Chemoenzymatic synthesis of human milk oligosaccharides (HMOs) takes full advantage of the advantages of high efficiency and strong selectivity of enzymatic synthesis, combined with the flexibility of chemical synthesis. CD BioGlyco has experienced chemical and biological scientists who provide you with high-level chemical and enzymatic synthesis services for HMO to meet the needs of various projects.

Advantages of Chemoenzymatic Synthesis for HMO

Chemical synthesis is a reliable method to obtain oligosaccharides with a definite structure. However, due to its numerous steps, strict operating conditions, and low overall yield, it is difficult to achieve the purpose of large-scale synthesis and use in the research of its biological functions. For another commonly used enzymatic synthesis, most of the enzymes come from mammals, which has problems such as low expression, narrow substrate adaptability, and complex separation and purification. The chemoenzymatic methods fully apply the advantages of high efficiency and selectivity of enzymatic synthesis and combine the flexibility of chemical synthesis, which realizes multiple effective combination modes, and has the characteristics of high efficiency, fast speed, high yield, product diversification, etc.

Chemoenzymatic strategies. Fig.1 Chemoenzymatic strategies. (Faijes, et al., 2019)

Classical Applications of Chemoenzymatic Synthesis of HMOs

At present, most of the confirmed structures of HMOs have a common core structure: lacto-N-tetraose (Galβ1-3GlcNAcβ1-3Galβ1-4Glc, LNT). It is synthesized by chemoenzymatic synthesis, that is the D-galactose-1-3-N-acetyl-D-hexosamine phosphorylase (BiGalHexNAcP) from Bifidobacterium infantis and the recombinant galactokinase (EcGalK) from Escherichia coliare used to catalyze a pot of two enzyme reaction to synthesize Galβ1-3G1cNAc disaccharide block. Then, with Galβ1-3G1cNAc disaccharide block and chemically synthesized lactose glycosides the core structure of HMO, LNT, was successfully and efficiently synthesized by chemical glycosylation.

As a substrate, LNT is further sialylated by using the "one pot multi-enzyme" synthesis system, Sialyllacto-N-tetraose a, (LSTa) has been successfully synthesized. As the core structure and key building block, LNT is also used as the substrate to synthesize many other sialylated or fucosylated HMOs, which needs to be further improved in the future.

HMO Production by chemoenzymatic Methods at CD BioGlyco

Due to the diversity and complexity of the products generated by the chemoenzymatic synthesis method, we also synthesize libraries including linear, biantennary, and triantennary glycans, etc.

The types of HMOs we synthesize by chemoenzymatic methods include but are not limited to LNT, LNnH, FLNnH I, FLNnH II, DFLNnH, etc, including some corresponding derivatives, bi-antennary HMO, and asymmetry multi-antennary HMO.

Abbreviation Full name
LNT Lacto-N-tetraose
LNT II Lacto-N-tetraose II
LNnH Lacto-N-neohexaose
FLNnH I, F-lacto-N-neohexaose I
FLNnH II F-lacto-N-neohexaose II
DFLNnH DF-lacto-N-neohexaose
3’SL 3’-Sialyllactose

CD BioGlyco has abundant enzymes for chemoenzymatic synthesis to complete the production of various HMO types, a small part of which is listed in the following table:

Enzyme Source
Galactokinase Bifidobacterium infantis
Pyrophosphorylase Arabidopsis thaliana
β1-4-N-galactosyltransferase Neisseria meningitides
Inorganic pyrophosphatase Escherichia coli
CMP-sialic acid synthetase Neisseria meningitides
α2, 6-sialyltransferase Photobacterium damselae
α1, 3-fuco-syltransferase Helicobacter pylori
α1, 2-fucosyltransferase Helicobacter mustelae

Advantages of Us

  • Produce a wide variety of structurally complex HMOs.
  • Realize regional selective protection.
  • High-yield synthesis.
  • Efficient reaction.

As an international leading biotechnology company, CD BioGlyco accumulates rich experience in chemoenzymatic methods of HMOs. Many of our authorities are willing to share their knowledge and experience in this field with you. If you are interested in the services we provide, please do not hesitate to contact us, we will reply in time.


  1. Faijes, M.; et al. Enzymatic and cell factory approaches to the production of human milk oligosaccharides. Biotechnology Advances. 2019, 37(5): 667-697.
This service is for Research Use Only, not intended for any clinical use.

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