Analysis of N-glycosylation of phospholipase A2 from venom of individual bees by microbore high-performance liquid chromatography-electrospray mass spectrometry using an ion trap mass spectrometer.

The N-linked oligosaccharides were released from the phospholipase A2 (PLA) with glycopeptidases and reductively aminated with the chromophore, p-aminobenzoic acid ethyl ester (ABEE). The ABEE-labeled oligosaccharides were separated by microbore high-performance liquid chromatography (micro-HPLC) using a reversed-phase column and analyzed by electrospray mass spectrometry. Differentiation between alpha-1,3 and alpha-1,6 core-fucosylated glycans was achieved by comparison the glycans released by glycopeptidases peptide-N-glycanase A (PNase A) and peptide-N-glycanase F (PNase F). All N-linked oligosaccharides except 3B and 3C could be identified in this approach. The analysis of PLA oligosaccharides from the venom of individual bees indicated that glycosylation patterns between the younger and the older bees were similar.