N- and <i>O</i>-glycosylation Analysis of Human C1-inhibitor Reveals Extensive Mucin-type <i>O</i>-Glycosylation.

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Stavenhagen K., Kayili H. M. , Holst S., Koeleman C. A. M. , Engel R., Wouters D., ...Daha Fazla

Molecular & cellular proteomics : MCP, cilt.17, ss.1225-1238, 2018 (SCI Expanded İndekslerine Giren Dergi) identifier identifier

  • Cilt numarası: 17 Konu: 6
  • Basım Tarihi: 2018
  • Doi Numarası: 10.1074/mcp.ra117.000240
  • Dergi Adı: Molecular & cellular proteomics : MCP
  • Sayfa Sayıları: ss.1225-1238


Human C1-inhibitor (C1-Inh) is a serine protease inhibitor and the major regulator of the contact activation pathway as well as the classical and lectin complement pathways. It is known to be a highly glycosylated plasma glycoprotein. However, both the structural features and biological role of C1-Inh glycosylation are largely unknown. Here, we performed for the first time an in-depth site-specific N- and O-glycosylation analysis of C1-Inh combining various mass spectrometric approaches, including C18-porous graphitized carbon (PGC)-LC-ESI-QTOF-MS/MS applying stepping-energy collision-induced dissociation (CID) and electron-transfer dissociation (ETD). Various proteases were applied, partly in combination with PNGase F and exoglycosidase treatment, in order to analyze the (glyco)peptides. The analysis revealed an extensively O-glycosylated N-terminal region. Five novel and five known O-glycosylation sites were identified, carrying mainly core1-type O-glycans. In addition, we detected a heavily O-glycosylated portion spanning from Thr(82)-Ser(121) with up to 16 O-glycans attached. Likewise, all known six N-glycosylation sites were covered and confirmed by this site-specific glycosylation analysis. The glycoforms were in accordance with results on released N-glycans by MALDI-TOF/TOF-MS/MS. The comprehensive characterization of C1-Inh glycosylation described in this study will form the basis for further functional studies on the role of these glycan modifications.