Microassay Analyses of Protein Glycosylation

The majority of secreted and cell surface proteins are glycosylated. To characterize accurately the carbohydrate moieties of oligosaccharide chains in glycosylated proteins, it is necessary to distinguish exactly which types of oligosaccharide are present and at which sites in the protein. The goal of the microassays presented here is to determine oligosaccharide structure and occupancy using lectin overlay assays. Lectins are multivalent binding proteins with the ability to bind specifically to certain sugar sequences in oligosaccharides and glycopeptides (). Lectin overlay assays take advantage of the ability of lectins to distinguish between different types of glycoproteins by virtue of their ability to recognize terminal sugars, thus allowing the chain type and peripheral antigenic components to be determined. In addition, the assay is important as it provides structural information of protein glycosylation prior to conformational analysis and identification of antigenically and biologically active oligosaccharides. The lectin overlay, in the form of a microassay, can be developed to give a high level of sensitivity for glycan detection. The specificities of the lectins in such assays appears to vary from previous studies using lectin affinity chromatography and hence must be characterized further