Glycosylation Site Analysis of Human Platelets by Electrostatic Repulsion Hydrophilic Interaction Chromatography

Clinical Proteomics

Katharina Lohrig¹  Dirk Wolters¹  Urs Lewandrowski²  René P. Zahedi²  Albert Sickmann² 

[1] AG Biomolecular Mass Spectrometry, Analytical Chemistry Department, Ruhr-University Bochum, Bochum, GermanyAG Biomolecular Mass Spectrometry, Analytical Chemistry Department, Ruhr-University Bochum, Bochum, GermanyAG Biomolecular Mass Spectrometry, Analytical Chemistry Department, Ruhr-University Bochum, Bochum, Germany;DFG-Research Center for Experimental Biomedicine, University Würzburg, Würzburg, GermanyDFG-Research Center for Experimental Biomedicine, University Würzburg, Würzburg, GermanyDFG-Research Center for Experimental Biomedicine, University Würzburg, Würzburg, Germany

关键词: Electrostatic repulsion hydrophilic interaction chromatography;    Glycosylation;    Mass spectrometry;    Platelet;   

DOI  :  10.1007/s12014-008-9006-z

来源: Humana Press Inc

【 摘 要 】

Abstract

Introduction

Glycosylations range among the most common posttranslational modifications with an estimated 50% of all proteins supposed to be glycosylated. These modifications are required for essential cellular processes including cell–cell recognition, protein structure and activity, e.g., of surface receptors, as well as subcellular localization of proteins. Beside the elucidation of the carbohydrate structures, the annotation of glycosylation sites is of primary interest as a basis for subsequent functional characterization. Although mass spectrometry is the method of choice for large-scale analysis of glycosylation sites, it requires initial enrichment of glycopeptides prior mass spectrometric detection in most cases.

Materials and Methods

In this paper, we present a novel approach for glycopeptide enrichment by electrostatic repulsion hydrophilic interaction chromatography (ERLIC). Glycopeptides were separated from the bulk of non-modified peptides and gradually eluted from the stationary phase with potential for isoform resolution. Applied to human platelets, 125 glycosylation sites on 66 proteins were identified including major platelet glycoproteins responsible for cellular function.

Conclusion

These sites add a major contribution to the now more than 250 glycosylation sites annotated for platelets, which enable the clinically relevant design of quantification assays for platelet glycoproteins.

【 授权许可】

Unknown