title: Polysaccharide microarrays for high-throughput screening of transglycosylase activities in plant extracts.
abstract: Polysaccharide transglycosylases catalyze disproportionation of polysaccharide molecules by cleaving glycosidic linkages in polysaccharide chains and transferring their cleaved portions to hydroxyl groups at the non-reducing ends of other polysaccharide or oligosaccharide molecules. In plant cell walls, transglycosylases have a potential to catalyze both cross-linking of polysaccharide molecules and grafting of newly arriving polysaccharide molecules into the cell wall structure during cell growth. Here we describe a polysaccharide microarray in form of a glycochip permitting simultaneous high-throughput monitoring of multiple transglycosylase activities in plant extracts. The glycochip, containing donor polysaccharides printed onto nitrocellulose-coated glass slides, was incubated with crude plant extracts, along with a series of fluorophore-labelled acceptor oligosaccharides. After removing unused labelled oligosaccharides by washing, fluorescence retained on the glycochip as a result of transglycosylase reaction was detected with a standard microarray scanner. The glycochip assay was used to detect transglycosylase activities in crude extracts from nasturtium (Tropaeolum majus) and mouse-ear cress (Arabidopsis thaliana). A number of previously unknown saccharide donor-acceptor pairs active in transglycosylation reactions that lead to the formation of homo- and hetero-glycosidic conjugates, were detected. Our data provide experimental support for the existence of diverse transglycosylase activities in crude plant extracts.
authors: Institute of Chemistry, Centre for Glycomics, Slovak Academy of Sciences, Dúbravská cesta 9, 84538, Bratislava, Slovakia.
authors: Ondrej
authors: O
authors: Kosík
authors: Richard P
authors: RP
authors: Auburn
authors: Steven
authors: S
authors: Russell
authors: Eva
authors: E
authors: Stratilová
authors: Sona
authors: S
authors: Garajová
authors: Maria
authors: M
authors: Hrmova
authors: Vladimír
authors: V
authors: Farkas
mesh_terms: D000327:Adsorption; D006031:Glycosylation; D006602:Hexosyltransferases / Q000378:metabolism*; D057166:High-Throughput Screening Assays / Q000379:methods*; D020128:Inhibitory Concentration 50; D046228:Microarray Analysis / Q000379:methods*; D010936:Plant Extracts / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D032388:Tropaeolum / Q000201:enzymology*
chemical_list: D010936:Plant Extracts; D011134:Polysaccharides; D006602:Hexosyltransferases