Glycan microarrays represent a high-throughput approach to determining the specificity of glycan-binding proteins against a large set of glycans in a single format. This chapter describes the use of a glycan microarray platform for evaluating the activity and substrate specificity of glycosyltransferases (GTs). The methodology allows simultaneous screening of hundreds of immobilized glycan acceptor substrates by in situ incubation of a GT and its appropriate donor substrate on the microarray surface. Using biotin-conjugated donor substrate enables direct detection of the incorporated sugar residues on acceptor substrates on the array. In addition, the feasibility of the method has been validated using label-free donor substrate combined with lectin-based detection of product to assess enzyme activity. Here, we describe the application of both procedures to assess the specificity of a recombinant human α2-6 sialyltransferase. This technique is readily adaptable to studying other glycosyltransferases. © Springer Science+Business Media New York 2013.
CITATION STYLE
Peng, W., Nycholat, C. M., & Razi, N. (2013). Glycan microarray screening assay for glycosyltransferase specificities. Methods in Molecular Biology, 1022, 1–14. https://doi.org/10.1007/978-1-62703-465-4_1
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