Alena Šedivá, Martina Zámorová , Dragana Robajac, Goran Miljuš, Miloš Šundericb Olgica Nedic, Jaroslav Katrlik
Institute of Chemistry, Slovak Academy of Sciences, Department of Glycobiotechnology, Dubravska cesta 9, 845 38 Bratislava, Slovakia
Glycosylation is a form of co-translational and post-translational modification of proteins. Glycans have a variety of structural and functional roles in membrane and secreted proteins. Changes in glycosylation are accompanied with changes in physiological state, which are also associated with various diseases such as cancer, AIDS, rheumatoid arthritis, multiple sclerosis and many others. Glycoprofiling thus could serve as a diagnostics tool for even early stage of diseases. A highly challenging is the fact that analytical detection platform for glycan analysis has to be able to detect very low levels of analytes. The aim of this work was to develop lectin-based protein microarray method for glycoprofiling of proteins and to apply this method for analyses of glycoproteins isolated from the samples by immunoprecipitation. We measured proteins and receptors of insulin-like growth factor (IGF) system isolated from serum, cytosol or membrane protein fraction of two groups of the samples, (1) serum of healthy persons and patients with colorectal cancer and healthy and tumor part of the tissue of the same patient, and (2) placental membranes from healthy mothers, mothers with pre-eclampsia, and mothers with diabetes. The microarray slides with spotted samples were incubated with set of biotinylated lectins with various glyco-specificity, labeled with streptavidin-fluorescent dye conjugate and scanned. The appropriateness of developed lectin-based protein microarray method for biosensing of changes in protein glycosylation of IGF system was evaluated, and the differences in protein glycosylation within each sample group (for colorectal cancer healthy vs. tumor tissue, for placental samples healthy vs. pre-eclampsia vs. diabetes) were quantified.
ACKNOWLEDGEMENT
This work is supported by the grants: VEGA 2/0162/14, APVV SK-SRB-2013-0028 and APVV 0282-11.