Isolation of Glycoproteins and Identification of Their N-Linked Glycosylation Sites (original) (raw)

Abstract

Protein glycosylation has long been recognized as a very common posttranslational modifi- cation. Protein glycosylation is prevalent in proteins destined for extracellular environments. These include proteins localized on the extracellular surface and those secreted to body fluids. In search of a method that has the potential to identify and quantify most proteins found in body fluids or the cell surface, we have recently developed a novel method for solid-phase extraction of formerly _N_-linked glycosylated peptides from glycoproteins. It has been shown that proteins secreted to body fluids or localized on the cell surface can be specifically enriched by this method. The technique is based on the conjugation of glycoproteins to a solid support using hydrazide chemistry, removal of nonglycosylated peptides by trypsin digestion, stable isotope labeling of glycopeptides, and the specific release of formerly _N_-linked glycosylated peptides via peptide-_N_-glycosidase. The recovered formerly _N_-linked glycopeptides are then identified and quantified by tandem mass spectrometry.

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Authors

  1. Hui Zhang
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  2. Ruedi Aebersold
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© 2006 Humana Press Inc.

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Zhang, H., Aebersold, R. (2006). Isolation of Glycoproteins and Identification of Their N-Linked Glycosylation Sites. In: New and Emerging Proteomic Techniques. Methods in Molecular Biology™, vol 328. Humana Press. https://doi.org/10.1385/1-59745-026-X:177

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