Detailed characterization of the O-linked glycosylation of the neuropilin-1 c/MAM-domain

作者：Windwarder Markus^*; Yelland Tamas; Djordjevic Snezana; Altmann Friedrich

来源：Glycoconjugate Journal, 2016, 33(3): 387-397.

DOI：10.1007/s10719-015-9602-x

• O-glycosylation
• neuropilin-1
• Electron-transfer dissociation
• MAM-domain
• O-GalNAc glycan

Neuropilins are involved in angiogenesis and neuronal development. The membrane proximal domain of neuropilin-1, called c or MAM domain based on its sequence conservation, has been implicated in neuropilin oligomerization required for its function. The c/MAM domain of human neuropilin-1 has been recombinantly expressed to allow for investigation of its propensity to engage in molecular interactions with other protein or carbohydrate components on a cell surface. We found that the c/MAM domain was heavily O-glycosylated with up to 24 monosaccharide units in the form of disialylated core 1 and core 2 O-glycans. Attachment sites were identified on the chymotryptic c/MAM peptide ETGATEKPTVIDSTIQSEFPTY by electron-transfer dissociation mass spectrometry (ETD-MS/MS). For highly glycosylated species consisting of carbohydrate to about 50 %, useful results could only be obtained upon partial desialylation. ETD-MS/MS revealed a hierarchical order of the initial O-GalNAc addition to the four different glycosylation sites. These findings enable future functional studies about the contribution of the described glycosylations in neuropilin-1 oligomerization and the binding to partner proteins as VEGF or galectin-1.
As a spin-off result the sialidase from Clostridium perfringens turned out to discriminate between galactose- and N-acetylgalactosamine-linked sialic acid.

• 出版日期2016-6