Screening for N-glycosylated proteins by liquid chromatography mass spectrometry.
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Bunkenborg J, Pilch BJ, Podtelejnikov AV, Wisniewski JR
Screening for N-glycosylated proteins by liquid chromatography mass spectrometry.
Proteomics. 2004 Feb;4(2):454-65.
- PubMed ID
- 14760718 [ View in PubMed]
- Abstract
In the last few years mass spectrometry has become the method of choice for characterization of post-translationally modified proteins. Whereas most protein chemical modifications are binary in the sense that only one change can be associated with a given residue, many different oligosaccharides can be attached to a glycosylation site residue. The detailed characterization of glycoproteins in complex biological samples is extremely challenging. However, information on N-glycosylation can be gained at an intermediary level. Here we demonstrate a procedure for mapping N-glycosylation sites in complex mixtures by reducing sample complexity and enriching glycoprotein content. Glycosylated proteins are selected by an initial lectin chromatography step and digested with endoproteinase Lys-C. Glycosylated peptides are then selected from the digest mixture by a second lectin chromatography step. The glycan components are removed with N-glycosidase F and the peptides digested with trypsin before analysis by on-line reversed-phase liquid chromatography mass spectrometry. Using two different lectins, concanavalin A and wheat germ agglutinin, this procedure was applied to human serum and a total of 86 N-glycosylation sites in 77 proteins were identified.
DrugBank Data that Cites this Article
- Polypeptides
Name UniProt ID Prothrombin P00734 Details Corticosteroid-binding globulin P08185 Details Antithrombin-III P01008 Details Transthyretin P02766 Details Serotransferrin P02787 Details Alpha-2-macroglobulin P01023 Details Alpha-1-acid glycoprotein 1 P02763 Details Serum paraoxonase/arylesterase 1 P27169 Details Alpha-1-antitrypsin P01009 Details Insulin-like growth factor-binding protein 3 P17936 Details Kininogen-1 P01042 Details von Willebrand factor P04275 Details Vitronectin P04004 Details Ceruloplasmin P00450 Details Fibronectin P02751 Details Fibrinogen gamma chain P02679 Details Plasma serine protease inhibitor P05154 Details Cholinesterase P06276 Details Plasma protease C1 inhibitor P05155 Details Beta-2-glycoprotein 1 P02749 Details Zinc-alpha-2-glycoprotein P25311 Details Complement C3 P01024 Details Carboxypeptidase N subunit 2 P22792 Details Alpha-1-acid glycoprotein 2 P19652 Details Ig alpha-2 chain C region P01877 Details Coagulation factor XII P00748 Details Complement C4-A P0C0L4 Details Complement C4-B P0C0L5 Details Apolipoprotein D P05090 Details Ig mu chain C region P01871 Details Afamin P43652 Details Alpha-1B-glycoprotein P04217 Details Alpha-2-HS-glycoprotein P02765 Details C4b-binding protein alpha chain P04003 Details Complement component C9 P02748 Details Complement factor H P08603 Details Complement factor I P05156 Details Clusterin P10909 Details Leucine-rich alpha-2-glycoprotein P02750 Details Lumican P51884 Details C4b-binding protein beta chain P20851 Details Inter-alpha-trypsin inhibitor heavy chain H1 P19827 Details Inter-alpha-trypsin inhibitor heavy chain H4 Q14624 Details Alpha-1-antichymotrypsin P01011 Details Apolipoprotein B-100 P04114 Details Hemopexin P02790 Details Haptoglobin P00738 Details