N-glycoproteomics of brain synapses and synaptic vesicles

At mammalian neuronal synapses, synaptic vesicle (SV) glycoproteins are essential for robust neurotransmission. Asparagine (N)-linked glycosylation is required for delivery of the major SV glycoproteins synaptophysin and SV2A to SVs. Despite this key role for N-glycosylation, the molecular compositions of SV N-glycans are largely unknown. In this study, we combined organelle isolation techniques and high-resolution mass spectrometry to characterize N-glycosylation at synapses and SVs from mouse brain. Detecting over 2,500 unique glycopeptides, we found that SVs harbor a distinct population of oligomannose and highly fucosylated N-glycans. Using complementary fluorescence methods, we identify at least one highly fucosylated N-glycan enriched in SVs compared with synaptosomes. High fucosylation was characteristic of SV proteins, plasma membrane proteins, and cell adhesion molecules with key roles in synaptic function and development. Our results define the N-glycoproteome of a specialized neuronal organelle and inform timely questions in the glycobiology of synaptic pruning and neuroinflammation.