Comprehensive site- and structure-specific profiling of N-glycosylation of edible bird's nest (EBN) proteome using label-free quantitative glycoproteomics.

Glycoproteins, which are involved in numerous biological functions, are among the most critical functional ingredients in an edible bird's nest (EBN). To gain a comprehensive understanding of the glycoprotein species within EBN, a label-free, site-specific glycoproteomic approach was used to analyze their N-glycoproteins, N-glycopeptides, and N-glycans systematically. A total of 127 N-glycoproteins were identified in EBN, of which 72 were found in house-EBN and 63 in cave-EBN, yielding 4195 and 5649 glycopeptides, respectively. Eight N-glycoproteins were common to both types, comprising 288 intact N-glycopeptides and 235 N-glycans. The results showed a relatively high abundance of terminally sialylated and core fucosylated N-glycans in EBN. Moreover, through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, it was observed that EBN N-glycoproteins predominantly participated in neurodegeneration-multiple illness, cell adhesion molecules, TNF signaling, and TGF-beta signaling pathways. These findings provide insights into EBN glycoprotein site-specific N-glycosylation and its biological roles and processes.