Screening, Isolation And Identification Of Human Glycan-binding Proteins

Glycans, as important informational moleculars, mediate various fundamental biological processes, including cell-cell recognition, immune response, cell adhension, cell growth and differentiation, et al. However, these important effects of carbohydrate can not perform independently. Only with their matching glycan-binding proteins (GBPs), can they accomplish their roles. To understand the biology behind protein-glycan interactions, it is imperative to identify all of the GBPs and the glycan ligands to which they bind.]n our study, carbohydrate microarray and biomagnetic separation techniques were used for the investigation of GBPs. And the experiment contains two parts:Part1:We used an integrated strategy based on carbohydrate microarrays and biomagnetic separation techniques to identify potential glycan-binding protein biomarkers. Sera samples from patients with hepatocellular carcinoma (HCC) were analyzed and compared with sera from healthy controls. The proteins were identified with LC-MS/MS and quantified with an emPAI method. Three glycans, fucose, Neu5Ac and lactose, were chosen based on the results of carbohydrate microarrays for the isolation of GBPs, respectively. In total, we identified:a. Fucose-binding proteins:49proteins in normal sera,45proteins in HCC sera; b. Neu5Ac-binding proteins:78proteins in normal sera,45proteins in HCC sera. c. Lactose-binding proteins:30proteins in normal sera,36proteins in HCC sera. This strategy could also be used in the discovery of new disease-specific biomarker for other diseases. Part2:Saliva from people of different ages and genders, were collected. And all the saliva, after labeled with Cy3, were analysed using a carbohydrate microarray which contains40glycans. The result of the microarrays were compared with each other to discover difference between ages and genders. For better interpretation of the data, glycans were divided into different classes according to their terminal monosaccharades. Surprisingly, similar trends were observed among glycans in the same class, which demonstrate the significance of terminal monosaccharade in glycan-GBP interactions. In additional, more changes were found among differnet ages than different genders. This research provided overall conditions of the GBPs in saliva of different groups of people, which might benefit the following research of saliva proteomics. However, more works are surely needed for the understanding of the physiological role of GBPs in saliva due to the complexity of saliva proteome and the limitation of carbohydrate microarrays.