Glycoarray Technologies: Deciphering Interactions from Proteins to Live Cell Responses

Abstract

Microarray technologies inspired the development of carbohydrate arrays. Initially, carbohydrate array technology was hindered by the complex structures of glycans and their structural variability. The first designs of glycoarrays focused on the HTP (high throughput) study of protein-glycan binding events, and subsequently more in-depth kinetic analysis of carbohydrate-protein interactions. However, the applications have rapidly expanded and now achieve successful discrimination of selective interactions between carbohydrates and, not only proteins, but also viruses, bacteria and eukaryotic cells, and most recently even live cell responses to immobilized glycans. Combining array technology with other HTP technologies such as mass spectrometry is expected to allow even more accurate and sensitive analysis. This review provides a broad overview of established glycoarray technologies (with a special focus on glycosaminoglycan applications) and their emerging applications to the study of complex interactions between glycans and whole living cells.

Keywords: glycobioarrays; glycoconjugates; glycomics; heparan sulfate; saccharide libraries.

Figure 1

Schematic diagram of the various applications of glycan arrays relating to applications including screening protein and antibody interactions with various glycans, virus and bacteria interactions with glycoproteins; and interactions of mammalian cells with glycosaminoglycan structures which lead to live cell responses relating to phosphorylation of cell signaling cascades. Symbols for glycan structures use nomenclature from [7]. Yellow circles: galactose; yellow squares: N-acetylgalactosamine; blue squares: N-acetylglucosamine; green circles: mannose; blue/white diamonds: glucuronic acid; brown/white diamond: iduronic acid; sulphation shown by 2S, 6S and NS; extra lines on the diamond represent unsaturated bonds.

Figure 2

Schematic diagram of a glycobioarray platform for screening live cell fibroblast growth factor signaling responses to immobilized heparin saccharides. See Puvirajesinghe et al. [39] for details. Saccharides immobilized onto an aminosilane glass surface via a Schiff’s base linkage with their reducing ends, is shown. Cells (shown using 40× magnification) can be overlaid onto the slide surface and cultured for a specified period, followed by fixation and immunostaining to detected specific epitopes for phosphorylation (green fluorescence for phosphorylated ERK and red fluorescence for total ERK) events using a microarray slide scanner. Symbols for glycan structures use nomenclature from [7]. Blue squares: N-acetylglucosamine; blue/white diamonds: glucuronic acid; brown/white diamond: iduronic acid; sulphation shown by 2S, 6S and NS; extra lines on the diamond represent unsaturated bonds.