Label-Free Analysis of O-glycosylation Site-Occupancy Based on the Signal Intensity of Glycopeptide/Peptide Ions

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  • Wada Yoshinao
    Department of Molecular Medicine, Osaka Medical Center and Research Institute for Maternal and Child Health

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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, apoliporotein 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.

Journal

  • Mass Spectrometry

    Mass Spectrometry 1 (2), A0008-A0008, 2012

    The Mass Spectrometry Society of Japan

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