Integrated GlycoProteome Analyzer (I-GPA) for Automated Identification and Quantitation of Site-Specific N-Glycosylation

Sci Rep. 2016 Feb 17;6:21175. doi: 10.1038/srep21175.

Abstract

Human glycoproteins exhibit enormous heterogeneity at each N-glycosite, but few studies have attempted to globally characterize the site-specific structural features. We have developed Integrated GlycoProteome Analyzer (I-GPA) including mapping system for complex N-glycoproteomes, which combines methods for tandem mass spectrometry with a database search and algorithmic suite. Using an N-glycopeptide database that we constructed, we created novel scoring algorithms with decoy glycopeptides, where 95 N-glycopeptides from standard α1-acid glycoprotein were identified with 0% false positives, giving the same results as manual validation. Additionally automated label-free quantitation method was first developed that utilizes the combined intensity of top three isotope peaks at three highest MS spectral points. The efficiency of I-GPA was demonstrated by automatically identifying 619 site-specific N-glycopeptides with FDR ≤ 1%, and simultaneously quantifying 598 N-glycopeptides, from human plasma samples that are known to contain highly glycosylated proteins. Thus, I-GPA platform could make a major breakthrough in high-throughput mapping of complex N-glycoproteomes, which can be applied to biomarker discovery and ongoing global human proteome project.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Algorithms
  • Automation, Laboratory
  • Blood Proteins / metabolism
  • Carcinoma, Hepatocellular / blood
  • Carcinoma, Hepatocellular / metabolism
  • Glycopeptides / chemistry
  • Glycopeptides / metabolism
  • Glycoproteins / chemistry
  • Glycoproteins / metabolism*
  • Glycosylation
  • Humans
  • Liver Neoplasms / blood
  • Liver Neoplasms / metabolism
  • Proteome
  • Proteomics / instrumentation
  • Proteomics / methods*
  • Reproducibility of Results

Substances

  • Blood Proteins
  • Glycopeptides
  • Glycoproteins
  • Proteome