Automated assignments of N- and O-site specific glycosylation with extensive glycan heterogeneity of glycoprotein mixtures

John S. Strum, Charles C. Nwosu, Serenus Hua, Scott R. Kronewitter, Richard R. Seipert, Robert J. Bachelor, Hyun Joo An, Carlito B Lebrilla

Research output: Contribution to journalArticle

51 Scopus citations

Abstract

Site-specific glycosylation (SSG) of glycoproteins remains a considerable challenge and limits further progress in the areas of proteomics and glycomics. Effective methods require new approaches in sample preparation, detection, and data analysis. While the field has advanced in sample preparation and detection, automated data analysis remains an important goal. A new bioinformatics approach implemented in software called GP Finder automatically distinguishes correct assignments from random matches and complements experimental techniques that are optimal for glycopeptides, including nonspecific proteolysis and high mass resolution liquid chromatography/tandem mass spectrometry (LC/MS/MS). SSG for multiple N- and O-glycosylation sites, including extensive glycan heterogeneity, was annotated for single proteins and protein mixtures with a 5% false-discovery rate, generating hundreds of nonrandom glycopeptide matches and demonstrating the proof-of-concept for a self-consistency scoring algorithm shown to be compliant with the target-decoy approach (TDA). The approach was further applied to a mixture of N-glycoproteins from unprocessed human milk and O-glycoproteins from very-low-density-lipoprotein (vLDL) particles.

Original languageEnglish (US)
Pages (from-to)5666-5675
Number of pages10
JournalAnalytical Chemistry
Volume85
Issue number12
DOIs
StatePublished - Jun 18 2013

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ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Strum, J. S., Nwosu, C. C., Hua, S., Kronewitter, S. R., Seipert, R. R., Bachelor, R. J., An, H. J., & Lebrilla, C. B. (2013). Automated assignments of N- and O-site specific glycosylation with extensive glycan heterogeneity of glycoprotein mixtures. Analytical Chemistry, 85(12), 5666-5675. https://doi.org/10.1021/ac4006556