Quantitative analysis of core fucosylation of serum proteins in liver diseases by LC-MS-MRM

J Proteomics. 2018 Oct 30;189:67-74. doi: 10.1016/j.jprot.2018.02.003. Epub 2018 Feb 7.

Abstract

Aberrant core fucosylation of proteins has been linked to liver diseases. In this study, we carried out multiple reaction monitoring (MRM) quantification of core fucosylated N-glycopeptides of serum proteins partially deglycosylated by a combination of endoglycosidases (endoF1, endoF2, and endoF3). To minimize variability associated with the preparatory steps, the analysis was performed without enrichment of glycopeptides or fractionation of serum besides the nanoRP chromatography. Specifically, we quantified core fucosylation of 22 N-glycopeptides derived from 17 proteins together with protein abundance of these glycoproteins in a cohort of 45 participants (15 disease-free control, 15 fibrosis and 15 cirrhosis patients) using a multiplex nanoUPLC-MS-MRM workflow. We find increased core fucosylation of 5 glycopeptides at the stage of liver fibrosis (i.e., N630 of serotransferrin, N107 of alpha-1-antitrypsin, N253 of plasma protease C1 inhibitor, N397 of ceruloplasmin, and N86 of vitronectin), increase of additional 6 glycopeptides at the stage of cirrhosis (i.e., N138 and N762 of ceruloplasmin, N354 of clusterin, N187 of hemopexin, N71 of immunoglobulin J chain, and N127 of lumican), while the degree of core fucosylation of 10 glycopeptides did not change. Interestingly, although we observe an increase in the core fucosylation at N86 of vitronectin in liver fibrosis, core fucosylation decreases on the N169 glycopeptide of the same protein. Our results demonstrate that the changes in core fucosylation are protein and site specific during the progression of fibrotic liver disease and independent of the changes in the quantity of N-glycoproteins. It is expected that the fully optimized multiplex LC-MS-MRM assay of core fucosylated glycopeptides will be useful for the serologic assessment of the fibrosis of liver. BIOLOGICAL SIGNIFICANCE: We have quantified the difference in core fucosylation among three comparison groups (healthy control, fibrosis and cirrhosis patients) using a sensitive and selective LC-MS-MRM method. Despite an overall increase in core fucosylation of many of the glycoproteins that we examined, core fucosylation changed in a protein- and site-specific manner. Moreover, increased and decreased fucosylation was observed on different N-glycopeptides of the same protein. Altered core fucosylation of N-glycopeptides might be used as an alternative serologic assay for the evaluation of fibrotic liver disease.

Keywords: Cirrhosis; Core fucosylation; Fibrosis; LC-MS-MRM; Serum.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antiviral Agents / therapeutic use
  • Blood Proteins / analysis*
  • Blood Proteins / metabolism*
  • Case-Control Studies
  • Chromatography, Liquid
  • Fucose / metabolism*
  • Glycosylation
  • Hepatitis C / blood
  • Hepatitis C / complications
  • Hepatitis C / drug therapy
  • Humans
  • Liver Cirrhosis / blood*
  • Liver Cirrhosis / metabolism
  • Randomized Controlled Trials as Topic
  • Tandem Mass Spectrometry / methods

Substances

  • Antiviral Agents
  • Blood Proteins
  • Fucose