LC-MS/MS characterization of O-glycosylation sites and glycan structures of human cerebrospinal fluid glycoproteins.

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Halim A, Ruetschi U, Larson G, Nilsson J

LC-MS/MS characterization of O-glycosylation sites and glycan structures of human cerebrospinal fluid glycoproteins.

J Proteome Res. 2013 Feb 1;12(2):573-84. doi: 10.1021/pr300963h. Epub 2013 Jan 11.

PubMed ID
23234360 [ View in PubMed
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Abstract

The GalNAc O-glycosylation on Ser/Thr residues of extracellular proteins has not been well characterized from a proteomics perspective. We previously reported a sialic acid capture-and-release protocol to enrich tryptic N- and O-glycopeptides from human cerebrospinal fluid glycoproteins using nano-LC-ESI-MS/MS with collision-induced dissociation (CID) for glycopeptide characterization. Here, we have introduced peptide N-glycosidase F (PNGase F) pretreatment of CSF samples to remove the N-glycans facilitating the selective characterization of O-glycopeptides and enabling the use of an automated CID-MS(2)/MS(3) search protocol for glycopeptide identification. We used electron-capture and -transfer dissociation (ECD/ETD) to pinpoint the glycosylation site(s) of the glycopeptides, identified as predominantly core-1-like HexHexNAc-O- structure attached to one to four Ser/Thr residues. We characterized 106 O-glycosylations and found Pro residues preferentially in the n - 1, n + 1, and/or n + 3 positions in relation to the Ser/Thr attachment site (n). The characterization of glycans and glycosylation sites in glycoproteins from human clinical samples provides a basis for future studies addressing the biological and diagnostic importance of specific protein glycosylations in relation to human disease.

DrugBank Data that Cites this Article

Polypeptides
NameUniProt ID
Neuron-specific vesicular protein calcyonQ9NYX4Details
Apolipoprotein A-IIP02652Details
Cathepsin DP07339Details
Apolipoprotein EP02649Details
Neuroendocrine convertase 1P29120Details
Plasma protease C1 inhibitorP05155Details
Neurocan core proteinO14594Details
Complement C4-AP0C0L4Details
Apolipoprotein C-IIP02655Details
Nucleobindin-1Q02818Details
Alpha-1-antichymotrypsinP01011Details
Amyloid-like protein 1P51693Details
Prostaglandin-H2 D-isomeraseP41222Details
HemopexinP02790Details