Label-free analysis of o-glycosylation site-occupancy based on the signal intensity of glycopeptide/peptide ions

Mass Spectrometry
Yoshinao Wada

Abstract

Mucin-type O-glycosylation is a major posttranslational modification of proteins. The level of O-glycosylation at a site could be useful in terms of evaluating various disease conditions. To address the feasibility of measuring O-glycosylation levels based on the glycopeptide ion intensity in a mass spectrum, apolipoprotein CIII (apoC3), a protein that contains a single core-1 O-glycan Gal-GalNAc disaccharide was analyzed by matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS). The intensity of protonated ions for an equimolar mixture of desialylated and deglycosylated apoC3s were the same in linear TOF measurements. No substantial in-source decay, including the cleavage of the protein-sugar linkage was observed. The glycopeptide derived from apoC3 and the unglycosylated counterpart, when analyzed by MALDI reflectron TOF MS indicated that post-source decay was minimal. These collective findings demonstrate the feasibility of label-free quantitation of O-glycan occupancy by MS when the glycans are small and neutral. This method provides a tool for use in glycoproteomics as a complement of our previous report (DOI: 10.1021/pr900913k) for calculating the saccharide composition of O-glycans.

References

Jun 3, 2005·Nature Reviews. Drug Discovery·Danielle H Dube, Carolyn R Bertozzi
Mar 7, 2006·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Yoshinao Wada
Sep 9, 2006·Cell·Kazuaki Ohtsubo, Jamey D Marth
May 18, 2007·Annual Review of Genomics and Human Genetics·Jaak Jaeken, Gert Matthijs
Jun 23, 2007·Current Molecular Medicine·Hudson H Freeze
Mar 27, 2009·Human Molecular Genetics·Vishwanathan HucthagowderZsolt Urban

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Citations

Jun 4, 2014·Mass Spectrometry Reviews·Yeong Hee AhnJong Shin Yoo
Jul 28, 2016·Journal of Pharmaceutical and Biomedical Analysis·Sachio YamamotoShigeo Suzuki
Apr 25, 2013·Anesthesiology·Matthias L Riess, Mihai V Podgoreanu

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