Abstract
The sites of mucin-type O-glycosylation are difficult to predict, making structural analysis by mass spectrometry indispensible. This chapter refers to state-of-the-art techniques in the site localization of O-linked glycans and their structural characterization in situ using tandem ESI and MALDI mass spectrometry. Detailed protocols are provided that describe the application of nano-LC-ESI-MS/MS with alternative fragmentation modes (collision-induced dissociation vs. electron-transfer dissociation) for the analysis of O-glycopeptides. Moreover, a top-down sequencing approach by MALDI-MS is presented that is based on the in-source decay of intact glycoproteins or large glycopeptides and allows a ladder sequencing of up to 70 amino acid residues from both termini with unequivocal assignment of modified sites.
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Acknowledgments
The structural work was supported in part by the Deutsche Forschungsgemeinschaft Grants HA 2092/15-1 and HA 2092/21-1 (to FGH).
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Hanisch, FG. (2012). O-Glycoproteomics: Site-Specifi c O-Glycoprotein Analysis by CID/ETD Electrospray Ionization Tandem Mass Spectrometry and Top-Down Glycoprotein Sequencing by In-Source Decay MALDI Mass Spectrometry. In: McGuckin, M., Thornton, D. (eds) Mucins. Methods in Molecular Biology, vol 842. Humana Press. https://doi.org/10.1007/978-1-61779-513-8_10
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