Research note: Structural identification of glycopeptides from chicken egg yolk protein

The structural characterization of glycopeptides is essential for elucidating their functional activity. In this study, the glycopeptide structures of chicken egg yolk proteins were identified comprehensively. Glycopeptides were obtained from egg yolk via trypsin digestion followed by hydrophilic interaction chromatography enrichment. Intact N- and O-glycopeptide structures of egg yolk proteins were analyzed using glycoproteomics techniques, and their potential functional activities were subsequently investigated. A total of 424 N-glycopeptides and 306 O-glycopeptides were identified, corresponding to 48 N-glycosites on 37 N-glycoproteins and 39 O-glycosites on 25 O-glycoproteins, respectively, demonstrating the extensive heterogeneity of glycosylation modifications. Twenty-two egg yolk glycoproteins were concurrently modified by N- and O-glycosylation. The identified glycopeptides exhibited diverse oligosaccharide chain compositions, demonstrating macro- and micro-heterogeneity. Apolipoprotein B yielded the most abundant glycopeptide structures, comprising 130 N-glycopeptides and 62 O-glycopeptides. N-glycoproteins were significantly enriched in immune-related signaling pathways, such as lysosome and regulation of actin cytoskeleton, whereas O-glycoproteins were significantly enriched in the spliceosome signaling pathway. These findings elucidated the structural characteristics of glycopeptides derived from egg yolk proteins and provided a theoretical basis for investigating their functional activities and potential applications as functional food ingredients.