Identification and quantification of N-linked glycoproteins using hydrazide chemistry, stable isotope labeling and mass spectrometry

Hui Zhang, Xiao jun Li, Daniel B. Martin, Ruedi Aebersold

Research output: Contribution to journalArticle

Abstract

Quantitative proteome profiling using stable isotope protein tagging and automated tandem mass spectrometry (MS/MS) is an emerging technology with great potential for the functional analysis of biological systems and for the detection of clinical diagnostic or prognostic marker proteins. Owing to the enormous complexity of proteomes, their comprehensive analysis is an as-yet-unresolved technical challenge. However, biologically or clinically important information can be obtained if specific, information-rich protein classes, or sub-proteomes, are isolated and analyzed. Glycosylation is the most common post-translational modification. Here we describe a method for the selective isolation, identification and quantification of peptides that contain N-linked carbohydrates. It is based on the conjugation of glycoproteins to a solid support using hydrazide chemistry, stable isotope labeling of glycopeptides and the specific release of formerly N-linked glycosylated peptides via peptide-N-glycosidase F (PNGase F). The recovered peptides are then identified and quantified by MS/MS. We applied the approach to the analysis of plasma membrane proteins and proteins contained in human blood serum.

Original languageEnglish (US)
Pages (from-to)660-666
Number of pages7
JournalNature Biotechnology
Volume21
Issue number6
DOIs
StatePublished - Jun 1 2003
Externally publishedYes

Fingerprint

Isotope Labeling
Glycoproteins
Isotopes
Labeling
Mass spectrometry
Mass Spectrometry
Proteome
Proteins
Peptides
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Glycosylation
Functional analysis
Glycopeptides
Biological systems
Cell membranes
Post Translational Protein Processing
Tandem Mass Spectrometry
Systems Analysis
Blood Proteins
Membrane Proteins

ASJC Scopus subject areas

  • Microbiology

Cite this

Identification and quantification of N-linked glycoproteins using hydrazide chemistry, stable isotope labeling and mass spectrometry. / Zhang, Hui; Li, Xiao jun; Martin, Daniel B.; Aebersold, Ruedi.

In: Nature Biotechnology, Vol. 21, No. 6, 01.06.2003, p. 660-666.

Research output: Contribution to journalArticle

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