Classification of Mucin-Type O -Glycopeptides Using Higher-Energy Collisional Dissociation in Mass Spectrometry.

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  • Additional Information
    • Source:
      Publisher: American Chemical Society Country of Publication: United States NLM ID: 0370536 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1520-6882 (Electronic) Linking ISSN: 00032700 NLM ISO Abbreviation: Anal Chem Subsets: MEDLINE
    • Publication Information:
      Original Publication: Washington, American Chemical Society.
    • Subject Terms:
      Glycopeptides/*chemistry ; Mass Spectrometry/*methods ; Urine/*chemistry; Databases, Factual ; Glycopeptides/classification ; Glycosylation ; Humans
    • Abstract:
      Changes in mucin-type O -glycosylation of human proteins affect protein function, immune response, and cancer progression. Since O -glycoproteins are characterized by the microheterogeneity of diverse O -glycans with no conserved sequence and the macroheterogeneity of multiple glycosylation sites on serine and/or threonine in human proteins, the assessment of different mucin types, such as Tn-antigen, core 1, and core 2, and their extended core types in O -glycopeptides, is extremely challenging. Here, we present an O-GlycoProteome Analyzer ( O -GPA) that automatically classifies mucin-type O -glycosylation using higher-energy collisional dissociation (HCD) in mass spectrometry. First, we estimated the number of GlcNAc residues using the intensity ratio of GlcNAc-specific fragment ions (HexNAc-CH 6 O 3 and HexNAc-2H 2 O) over GalNAc-specific fragment ions (HexNAc-C 2 H 6 O 3 and HexNAc-C 2 H 4 O 2 ) in the HCD spectrum. Furthermore, we classified the different mucin types of O -glycopeptides from characteristic B 2 (HexNAc 2 ) or Y 2α (PEP + HexNAc 2 ), and Y 2β (PEP + HexNAcHex) fragment ions, along with the number of GlcNAc. Furthermore, O -GPA automatically determined single or multiple O -glycosylation, regardless of the mucin types. The mucin type of O -glycopeptides from human urine and plasma was confirmed with an overall accuracy of 96%. We found 97 core 1, 56 core 2, 13 extended core 1, and 12 extended core 2 glycopeptides from urine; and 22 core 1, 13 core 2, 7 extended core 1, 1 extended core 2, and 1 Tn-antigen from plasma. Our strategy can be used to successfully characterize specific mucin types of O -glycoproteins in human biological samples.
    • Accession Number:
      0 (Glycopeptides)
    • Publication Date:
      Date Created: 20200626 Date Completed: 20210218 Latest Revision: 20210218
    • Publication Date:
      20250114
    • Accession Number:
      10.1021/acs.analchem.0c01218
    • Accession Number:
      32584546