| Literature DB >> 34626371 |
Fredrik Noborn1, Mahnaz Nikpour1, Andrea Persson1, Carina Sihlbom2, Jonas Nilsson1,3, Göran Larson4,5.
Abstract
In this chapter, we describe a glycoproteomic approach for the identification of novel chondroitin sulfate proteoglycans (CSPGs) using a combination of biochemical enrichments, enzymatic digestions, and nanoscale liquid chromatography tandem mass spectrometry (nLC-MS/MS) analysis. The identification is achieved by trypsin digestion of CSPG-containing samples, followed by enrichment of chondroitin sulfate (CS) glycopeptides by strong anion exchange chromatography (SAX). The enriched CS glycopeptides are then digested with chondroitinase ABC to depolymerize the CS polysaccharides, generating a residual hexasaccharide structure, composed of the linkage region tetrasaccharide extended with a terminal dehydrated disaccharide, still attached to the peptide. The obtained CS glycopeptides are analyzed by nLC-MS/MS, and the generated data sets are evaluated through proteomic software with adjustment in the settings to allow for glycopeptide identification. This approach has enabled the identification of several novel core proteins in human samples and in Caenorhabditis elegans. Here we specifically describe the procedure for the enrichment and characterization of CS glycopeptides from human cerebrospinal fluid (CSF).Entities:
Keywords: Chondroitin sulfate; Core proteins; Glycopeptides; Glycoproteomics; Heparan sulfate; Higher-energy collision dissociation; Linkage region; Proteoglycans; Tandem mass spectrometry
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Year: 2022 PMID： 34626371 DOI： 10.1007/978-1-0716-1398-6_7
Source DB: PubMed Journal: Methods Mol Biol ISSN： 1064-3745