title: Characterization of fibrinogen glycosylation and its importance for serum/plasma N-glycome analysis.
abstract: The majority of proteins present in human serum/plasma are glycoproteins, validating this fluid as an ideal starting material for N-glycan analysis and discovery of potential biomarkers. The glycoprotein content for both serum and plasma is very similar, except for proteins removed in the coagulation process, including fibrinogen. Our aim was to characterize fibrinogen glycosylation in order to determine its contribution to differences between serum and plasma N-glycomes. N-Glycans from human fibrinogen were released, labeled, and analyzed by HILIC-HPLC and MS. Structural characterization of fibrinogen subunits revealed that the α chain was not N-glycosylated, whereas β and γ contained identical oligosaccharide structures, mainly biantennary digalactosylated monosialylated structures (A2G2S1) and biantennary digalactosylated disialylated structures (A2G2S2). Blood was collected from five healthy volunteers into four testing tubes: silicone-coated glass for serum and EDTA, Na-heparin, and Li-heparin glass tubes for plasma. N-Glycans were analyzed using the high-throughput HILIC-HPLC method. N-Glycan profiles from serum and plasma samples differed largely in glycans identified in fibrinogen, suggesting that this glycoprotein represents a major factor distinguishing these body fluids. This result emphasizes the important of consistent body fluid collection practices in biomarker discovery studies.
authors.affiliation: NIBRT Dublin-Oxford Glycobiology Laboratory, National Institute for Bioprocessing Research and Training, Fosters Avenue, Mount Merrion, Blackrock, Co Dublin, Ireland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Adamczyk
authors.forename: Weston B
authors.initials: WB
authors.lastname: Struwe
authors.forename: Altan
authors.initials: A
authors.lastname: Ercan
authors.forename: Peter A
authors.initials: PA
authors.lastname: Nigrovic
authors.forename: Pauline M
authors.initials: PM
authors.lastname: Rudd
mesh_terms: D001798:Blood Proteins / Q000737:chemistry / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D005340:Fibrinogen / Q000031:analogs & derivatives* / Q000737:chemistry / Q000302:isolation & purification; D006023:Glycoproteins / Q000097:blood / Q000737:chemistry / Q000302:isolation & purification; D006031:Glycosylation; D006801:Humans; D009844:Oligosaccharides / Q000737:chemistry; D011134:Polysaccharides / Q000097:blood / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D001798:Blood Proteins; D006023:Glycoproteins; D009844:Oligosaccharides; D011134:Polysaccharides; C044548:glycosylated fibrinogen; D005340:Fibrinogen