Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry.

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Liu T, Qian WJ, Gritsenko MA, Camp DG 2nd, Monroe ME, Moore RJ, Smith RD

Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry.

J Proteome Res. 2005 Nov-Dec;4(6):2070-80.

PubMed ID
16335952 [ View in PubMed
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Abstract

The enormous complexity, wide dynamic range of relative protein abundances of interest (over 10 orders of magnitude), and tremendous heterogeneity (due to post-translational modifications, such as glycosylation) of the human blood plasma proteome severely challenge the capabilities of existing analytical methodologies. Here, we describe an approach for broad analysis of human plasma N-glycoproteins using a combination of immunoaffinity subtraction and glycoprotein capture to reduce both the protein concentration range and the overall sample complexity. Six high-abundance plasma proteins were simultaneously removed using a pre-packed, immobilized antibody column. N-linked glycoproteins were then captured from the depleted plasma using hydrazide resin and enzymatically digested, and the bound N-linked glycopeptides were released using peptide-N-glycosidase F (PNGase F). Following strong cation exchange (SCX) fractionation, the deglycosylated peptides were analyzed by reversed-phase capillary liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Using stringent criteria, a total of 2053 different N-glycopeptides were confidently identified, covering 303 nonredundant N-glycoproteins. This enrichment strategy significantly improved detection and enabled identification of a number of low-abundance proteins, exemplified by interleukin-1 receptor antagonist protein (approximately 200 pg/mL), cathepsin L (approximately 1 ng/mL), and transforming growth factor beta 1 (approximately 2 ng/mL). A total of 639 N-glycosylation sites were identified, and the overall high accuracy of these glycosylation site assignments as assessed by accurate mass measurement using high-resolution liquid chromatography coupled to Fourier transform ion cyclotron resonance mass spectrometry (LC-FTICR) is initially demonstrated.

DrugBank Data that Cites this Article

Polypeptides
NameUniProt ID
Coagulation factor XIII A chainP00488Details
Coagulation factor VIIIP00451Details
Vascular endothelial growth factor receptor 3P35916Details
Coagulation factor VP12259Details
ProthrombinP00734Details
Carcinoembryonic antigen-related cell adhesion molecule 1P13688Details
Dopamine beta-hydroxylaseP09172Details
Corticosteroid-binding globulinP08185Details
Angiotensin-converting enzymeP12821Details
Transcobalamin-1P20061Details
Antithrombin-IIIP01008Details
TransthyretinP02766Details
Vitamin K-dependent protein CP04070Details
Integrin alpha-IIbP08514Details
Mast/stem cell growth factor receptor KitP10721Details
Low-density lipoprotein receptorP01130Details
SerotransferrinP02787Details
Thyroxine-binding globulinP05543Details
Fibrinogen alpha chainP02671Details
Sex hormone-binding globulinP04278Details
Voltage-dependent calcium channel subunit alpha-2/delta-1P54289Details
Heparin cofactor 2P05546Details
Macrophage colony-stimulating factor 1 receptorP07333Details
Dipeptidyl peptidase 4P27487Details
Complement C5P01031Details
Lysosomal alpha-glucosidaseP10253Details
Alpha-2-macroglobulinP01023Details
Alpha-1-acid glycoprotein 1P02763Details
Serum paraoxonase/arylesterase 1P27169Details
Cathepsin DP07339Details
Vitamin K-dependent protein SP07225Details
Alpha-1-antitrypsinP01009Details
Tissue factor pathway inhibitorP10646Details
Insulin-like growth factor-binding protein 3P17936Details
Beta-glucuronidaseP08236Details
Transforming growth factor beta-1P01137Details
Membrane primary amine oxidaseQ16853Details
Kininogen-1P01042Details
Coagulation factor XIP03951Details
Basement membrane-specific heparan sulfate proteoglycan core proteinP98160Details
Integrin beta-2P05107Details
AngiotensinogenP01019Details
VitronectinP04004Details
MyeloperoxidaseP05164Details
Aminopeptidase NP15144Details
Neutrophil gelatinase-associated lipocalinP80188Details
Intercellular adhesion molecule 1P05362Details
CD44 antigenP16070Details
Vascular cell adhesion protein 1P19320Details
Alpha-2-antiplasminP08697Details
CeruloplasminP00450Details
Integrin beta-3P05106Details
FibronectinP02751Details
Serum amyloid P-componentP02743Details
ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase 2Q10588Details
Complement C1s subcomponentP09871Details
Complement factor BP00751Details
Fibrinogen beta chainP02675Details
Fibrinogen gamma chainP02679Details
Complement C1r subcomponentP00736Details
Complement C1q subcomponent subunit AP02745Details
Laminin subunit beta-1P07942Details
Laminin subunit gamma-1P11047Details
Prolow-density lipoprotein receptor-related protein 1Q07954Details
Plasma serine protease inhibitorP05154Details
Asialoglycoprotein receptor 2P07307Details
Neuropilin-1O14786Details
Fibroblast growth factor receptor 1P11362Details
Cathepsin L1P07711Details
CholinesteraseP06276Details
Glutamyl aminopeptidaseQ07075Details
Plasma kallikreinP03952Details
Plasma protease C1 inhibitorP05155Details
Beta-2-glycoprotein 1P02749Details
Zinc-alpha-2-glycoproteinP25311Details
Integrin beta-4P16144Details
Scavenger receptor cysteine-rich type 1 protein M130Q86VB7Details
Leptin receptorP48357Details
Amyloid beta A4 proteinP05067Details
Cadherin-5P33151Details
Carboxypeptidase B2Q96IY4Details
Complement C3P01024Details
Carboxypeptidase N subunit 2P22792Details
Alpha-mannosidase 2Q16706Details
Prolyl endopeptidase FAPQ12884Details
Cholesteryl ester transfer proteinP11597Details
Monocyte differentiation antigen CD14P08571Details
Serum paraoxonase/lactonase 3Q15166Details
Receptor-type tyrosine-protein phosphatase SQ13332Details
Integrin beta-1P05556Details
BDNF/NT-3 growth factors receptorQ16620Details
Carbonic anhydrase-related protein 11O75493Details
Alpha-1-acid glycoprotein 2P19652Details
Aggrecan core proteinP16112Details
Apolipoprotein MO95445Details
Poliovirus receptorP15151Details
Tyrosine-protein kinase MerQ12866Details
N-acetylmuramoyl-L-alanine amidaseQ96PD5Details
Carboxypeptidase QQ9Y646Details
Coagulation factor XIIP00748Details
Complement C4-AP0C0L4Details
Interleukin-1 receptor antagonist proteinP18510Details
Transferrin receptor protein 1P02786Details
Apolipoprotein DP05090Details
Selenoprotein PP49908Details
AfaminP43652Details
Neural cell adhesion molecule L1P32004Details
Macrophage receptor MARCOQ9UEW3Details
Alpha-1B-glycoproteinP04217Details
Alpha-2-HS-glycoproteinP02765Details
C4b-binding protein alpha chainP04003Details
Complement component C8 beta chainP07358Details
Complement component C9P02748Details
Complement factor HP08603Details
Complement factor IP05156Details
ClusterinP10909Details
Insulin-like growth factor-binding protein complex acid labile subunitP35858Details
Inter-alpha-trypsin inhibitor heavy chain H2P19823Details
Leucine-rich alpha-2-glycoproteinP02750Details
LumicanP51884Details
KallistatinP29622Details
Pigment epithelium-derived factorP36955Details
Apolipoprotein L1O14791Details
C4b-binding protein beta chainP20851Details
Complement component C8 alpha chainP07357Details
Coagulation factor XIII B chainP05160Details
Ficolin-3O75636Details
Inter-alpha-trypsin inhibitor heavy chain H3Q06033Details
Inter-alpha-trypsin inhibitor heavy chain H4Q14624Details
Immunoglobulin J chainP01591Details
Pregnancy zone proteinP20742Details
Alpha-1-antichymotrypsinP01011Details
A disintegrin and metalloproteinase with thrombospondin motifs 13Q76LX8Details
UDP-glucuronosyltransferase 2B10P36537Details
Phospholipid transfer proteinP55058Details
Lysophosphatidic acid receptor 4Q99677Details
Ectonucleoside triphosphate diphosphohydrolase 5O75356Details
Prostaglandin-H2 D-isomeraseP41222Details
Apolipoprotein B-100P04114Details
HemopexinP02790Details
Histidine-rich glycoproteinP04196Details
HaptoglobinP00738Details
Interleukin-6 receptor subunit betaP40189Details
Immunoglobulin heavy constant gamma 3P01860Details
Angiopoietin-related protein 3Q9Y5C1Details
Lymphatic vessel endothelial hyaluronic acid receptor 1Q9Y5Y7Details
Extracellular superoxide dismutase [Cu-Zn]P08294Details