Labeling of Mucin-Type O -Glycans for Quantification Using Liquid Chromatography and Fluorescence Detection

作者：Marc Safferthal, Leïla Bechtella, Andreas Zappe, Gaël M. Vos, Kevin Pagel · 发表于：ACS Measurement Science Au · 年份：2024 · DOI：10.1021/acsmeasuresciau.3c00071 · 被引用次数：15 · 研究领域：Glycosylation and Glycoproteins Research、Carbohydrate Chemistry and Synthesis、Proteoglycans and glycosaminoglycans research

High Resolution Image Download MS PowerPoint Slide O -glycosylation is a common post-translational modification that is essential for the defensive properties of mucus barriers. Incomplete and altered O -glycosylation is often linked to severe diseases, such as cancer, cystic fibrosis, and chronic obstructive pulmonary disease. Originating from a nontemplate-driven biosynthesis, mucin-type O -glycan structures are very complex. They are often present as heterogeneous mixtures containing multiple isomers. Therefore, the analysis of complex O -glycan mixtures usually requires hyphenation of orthogonal techniques such as liquid chromatography (LC), ion mobility spectrometry, and mass spectrometry (MS). However, MS-based techniques are mainly qualitative. Moreover, LC separation of O -glycans often lacks reproducibility and requires sophisticated data treatment and analysis. Here we present a mucin-type O -glycomics analysis workflow that utilizes hydrophilic interaction liquid chromatography for separation and fluorescence labeling for detection and quantification. In combination with mass spectrometry, a detailed analysis on the relative abundance of specific mucin-type O -glycan compositions and features, such as fucose, sialic acids, and sulfates, is performed. Furthermore, the average number of monosaccharide units of O -glycans in different samples was determined. To demonstrate universal applicability, the method was tested on mucins from different tissue types and mammals...