New Methods for the Characterization of Human Glycoproteins by LC-MS/MStechniques

Abstract

The glycosylation of proteins has numerous important functions in nature. Next to the stability and solubility of proteins, glycosylation is relevant for cellular recognition processes and immune modulation. Changes in glycosylation reflect the physiological and pathological state of a cell or an organism. A sophisticated structural characterization of glycans is needed to understand their biological relevance, but also for the development and quality control of biopharmaceuticals or for the development of glycan-based biomarkers. Mass spectrometry has become a well-established technique for the structural analysis of protein modifications due to low detection limits and high mass accuracy. In combination with other analytical methods such as liquid chromatography, mass spectrometry has made it possible to analyze complex glycosylation despite their structural complexity and diversity. Nevertheless, the development of new analytical approaches and MS-based techniques as well as their validation is necessary to further improve the analysis of glycans. This applies in particular to the quantification of glycans, the detection of low abundant structures, and the implementation of robust and timely methods. The aim of this work was therefore to evaluate different mass spectrometric methods for the structural characterization of glycoproteins and to analyze the glycosylation of different plasma proteins from several human plasma samples.

The glycosylation of proteins has numerous important functions in nature. Next to the stability and solubility of proteins, glycosylation is relevant for cellular recognition processes and immune modulation. Changes in glycosylation reflect the physiological and pathological state of a cell or an organism. A sophisticated structural characterization of glycans is needed to understand their biological relevance, but also for the development and quality control of biopharmaceuticals or for the development of glycan-based biomarkers. Mass spectrometry has become a well-established technique for the structural analysis of protein modifications due to low detection limits and high mass accuracy. In combination with other analytical methods such as liquid chromatography, mass spectrometry has made it possible to analyze complex glycosylation despite their structural complexity and diversity. Nevertheless, the development of new analytical approaches and MS-based techniques as well as their validation is necessary to further improve the analysis of glycans. This applies in particular to the quantification of glycans, the detection of low abundant structures, and the implementation of robust and timely methods. The aim of this work was therefore to evaluate different mass spectrometric methods for the structural characterization of glycoproteins and to analyze the glycosylation of different plasma proteins from several human plasma samples.