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One-pot three-enzyme chemoenzymatic approach to the synthesis of sialosides containing natural and non-natural functionalities

Nature Protocols volume 1, pages 2485–2492 (2006)Cite this article

Abstract

Chemoenzymatic synthesis, which combines the flexibility of chemical synthesis and the high selectivity of enzymatic synthesis, is a powerful approach to obtain complex carbohydrates. It is a preferred method for synthesizing sialic acid-containing structures, including those with diverse naturally occurring and non-natural sialic acid forms, different sialyl linkages and different glycans that link to the sialic acid. Starting from N-acetylmannosamine, mannose or their chemically or enzymatically modified derivatives, sialic acid aldolase-catalyzed condensation reaction leads to the formation of sialic acids and their derivatives. These compounds are subsequently activated by a CMP-sialic acid synthetase and transferred to a wide range of suitable acceptors by a suitable sialyltransferase for the formation of sialosides containing natural and non-natural functionalities. The three-enzyme coupled synthesis of sialosides can be carried out in one pot without the isolation of intermediates. The time for synthesis is 4–18 h. Purification and characterization of the product can be completed within 2–3 d.

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Figure 1
Figure 2: Schematic presentation of the one-pot three-enzyme chemoenzymatic approach to the synthesis of sialosides containing natural and non-natural functionalities.
Figure 3: Applications of the one-pot three-enzyme system to modify glycoconjugates or cells containing a terminal galactose or N-acetylgalactosamine residue.
Figure 4
Figure 5: TLC analysis of one-pot three-enzyme synthesis of sialosides using ManNAc as a sialic acid precursor and LacβProN3 as an acceptor substrate.

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Acknowledgements

This work was supported by the Mizutani Foundation for Glycoscience, NIH R01GM076360, and start-up funds from the Regents of the University of California.

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

    Hai Yu, Harshal A. Chokhawala, Shengshu Huang & Xi Chen

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Correspondence to Xi Chen.

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Yu, H., Chokhawala, H., Huang, S. et al. One-pot three-enzyme chemoenzymatic approach to the synthesis of sialosides containing natural and non-natural functionalities. Nat Protoc 1, 2485–2492 (2006). https://doi.org/10.1038/nprot.2006.401

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