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Oligosaccharide microarrays for high-throughput detection and specificity assignments of carbohydrate-protein interactions

Nature Biotechnology volume 20, pages 1011–1017 (2002)Cite this article

Abstract

We describe microarrays of oligosaccharides as neoglycolipids and their robust display on nitrocellulose. The arrays are sourced from glycoproteins, glycolipids, proteoglycans, polysaccharides, whole organs, or from chemically synthesized oligosaccharides. We show that carbohydrate-recognizing proteins single out their ligands not only in arrays of homogeneous oligosaccharides but also in arrays of heterogeneous oligosaccharides. Initial applications have revealed new findings, including: (i) among O-glycans in brain, a relative abundance of the Lewisx sequence based on N-acetyllactosamine recognized by anti-L5, and a paucity of the Lewisx sequence based on poly-N-acetyllactosamine recognized by anti-SSEA-1; (ii) insights into chondroitin sulfate oligosaccharides recognized by an antiserum and an antibody (CS-56) to chondroitin sulfates; and (iii) binding of the cytokine interferon-γ (IFN-γ) and the chemokine RANTES to sulfated sequences such as HNK-1, sulfo-Lewisx, and sulfo-Lewisa, in addition to glycosaminoglycans. The approach opens the way for discovering new carbohydrate-recognizing proteins in the proteome and for mapping the repertoire of carbohydrate recognition structures in the glycome.

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Figure 1: Immunological detection of oligosaccharides immobilized as NGLs on nitrocellulose and PVDF membranes.
Figure 2: Detection of epitope or ligand-bearing oligosaccharides on nitrocellulose membranes probed with carbohydrate-recognizing proteins.
Figure 3: Surveying the features of antigen-positive oligosaccharides within arrays derived from chondroitin sulfates A, B, and C.
Figure 4: Detection of the selective expression of carbohydrate differentiation antigens in brain O-glycan arrays.
Figure 5: Deconvolution by TLC, antibody binding, and mass spectrometry to examine the features of two distinct Lewisx -bearing oligosaccharides.

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Acknowledgements

This work is supported by a program grant G9601454 from the UK Medical Research Council. We thank D. Wang and W. Loveless for stimulating discussions, C. Herbert for technical support, and C. Kelly for assistance in compiling the manuscript.

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  1. Shigeyuki Fukui: Department of Biotechnology, Faculty of Engineering, Kyoto Sangyo University, Kyoto, Japan (on leave).

Authors and Affiliations

  1. Glycosciences Laboratory, Imperial College Faculty of Medicine, Northwick Park Hospital, Watford Road, Harrow, HA1 3UJ, Middlesex, UK

    Shigeyuki Fukui, Ten Feizi, Christine Galustian, Alexander M. Lawson & Wengang Chai

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Correspondence to Ten Feizi.

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Fukui, S., Feizi, T., Galustian, C. et al. Oligosaccharide microarrays for high-throughput detection and specificity assignments of carbohydrate-protein interactions. Nat Biotechnol 20, 1011–1017 (2002). https://doi.org/10.1038/nbt735

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