Macquarie U - Glycan Isomer Discrimination Product Ions

Discrimination of Isomers of Released N- and O-Glycans Using Diagnostic Product Ions in Negative Ion PGC-LC-ESI-MS/MS
  • Organism: Sus scrofa, Bos taurus, Homo sapiens
  • Instrument: Velos Plus,HCTultra
  • SpikeIn: No
  • Keywords: Diagnostic ions; Glycomics; MS/MS; Skyline
  • Submitter: Christopher Ashwood
Profiling cellular protein glycosylation is challenging due to the presence of highly similar glycan structures that play diverse roles in cellular physiology. As the anomericity and the exact linkage type of a single glycosidic bond can influence glycan function, there is a demand for improved and automated methods to confirm detailed structural features and to discriminate between structurally similar isomers, overcoming a significant bottleneck in the analysis of data generated by glycomics experiments. We used porous graphitized carbon-LC-ESI-MS/MS to separate and detect released N- and O-glycan isomers from mammalian model glycoproteins using negative mode resonance activation CID-MS/MS. By interrogating similar fragment spectra from closely related glycan isomers that differ only in arm position and sialyl linkage, product fragment ions for discrimination between these features were discovered. Using the Skyline software, at least two diagnostic fragment ions of high specificity were validated for automated discrimination of sialylation and arm position in N-glycan structures, and sialylation in O-glycan structures, complementing existing structural diagnostic ions. These diagnostic ions were shown to be useful for isomer discrimination using both linear and 3D ion trap mass spectrometers when analyzing complex glycan mixtures from cell lysates. Skyline was found to serve as a useful tool for automated assessment of glycan isomer discrimination. This platform-independent workflow can potentially be extended to automate the characterization and quantitation of other challenging glycan isomers
Experiment Description
Identification and validation of diagnostic MS2-based product ions for discriminating between glycan isomers.
Sample Description
N- and O-glycans released from purified standards.
Created on 8/17/18, 11:11 AM
Clustergrammer Heatmap
Download 10:01:1312281 10:01:0212262 10:00:5012282 10:00:3812262 10:00:2812261 10:00:14155171 09:59:5212281