The new strategy is revolutionizing glycoproteomics research

Protein glycosylation is a basic biological process regulating various cellular functions, but its study is hampered by the complexity and low abundance of glycoproteins. In a groundbreaking study published in , scientists from Fudan University developed a chemical ligation-based glycopeptide enrichment strategy called HG-TC. This method uses advanced solid-phase materials and bioorthogonal chemistry to simultaneously identify multiple types of glycosylations, including N-glycosites, O-GlcNAc and O-GalNAc sites, as well as N-glycans (see image below). This HG-TC strategy allows the enrichment of glycopeptides via azide-alkyne cycloaddition reactions and their release via trypsin cleavage. This single-tube workflow minimizes sample loss while maintaining high repeatability.

This approach provides a time-efficient workflow with exceptional scalability, enabling the identification of over 900 O-GlcNAc sites and 800 N-glycosites in HeLa cells in a single experiment, using minimal sample amounts. The team noted that “So”

The research team also applied this strategy to analyze HeLa cell samples under oxidative stress. They found distinct spatial patterns of glycosylation between the nucleus and cytoplasm, providing valuable information on the dynamic role of glycosylation in cellular responses. This study provides a robust tool for glycoproteomics research, helping to understand the dynamic role of glycosylation in cell signaling and disease mechanisms. For glycoproteomics researchers, this method represents a significant step forward, simplifying complex workflows while maintaining high data quality. The findings are particularly important in studying glycosylation changes in cancer and other diseases.