Macquarie U - Glycan Isomer Discrimination Product Ions

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
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
Flag FileDownloadCreatedListsSmall MoleculesPrecursorsTransitionsReplicates (121 MB)2019-06-139001922,76496 (12 MB)2019-06-1081016283018 (5 MB)2019-06-108101628548 (81 MB)2019-06-109001922,76266 (140 MB)2019-06-089001922,76496 (110 MB)2019-06-089001922,76491 (100 MB)2019-06-079001922,75866 (7 MB)2019-06-04802417496 (8 MB)2019-06-04802418396 (7 MB)2019-06-03802316196 (140 MB)2019-06-039001922,76496 (7 MB)2019-06-03802416596
all (46 MB)2019-05-311305268,5337
20180518 multiplex quantification - APOA4 only (265 MB)2019-05-311064864
20180518 multiplex quantification only (380 MB)2019-05-312807050268 (120 MB)2019-05-3116028684820
500 µE (PG-500uE_2ul_2019-05-16_15-23-09) (7 MB)2019-05-2927010555811 (3 MB)2019-05-292001225758 (13 MB)2019-05-293101366528
200µE (R5-8_V2_2019-05-16_15-31-29) (8 MB)2019-05-292809751616 (1 GB)2019-05-283,228027,28985,17542 (1 GB)2019-05-243,731036,39395,63942 (162 MB)2019-05-222009862412 (531 MB)2019-05-224,007016,22947,85630 (1 GB)2019-05-223,482030,60290,48842
Janschitz et al_PRM_results_Panorama (611 MB)2019-05-09405833712 (16 MB)2019-04-2981016287833 (43 MB)2019-04-2690019292496 (128 MB)2019-04-269001921,00296 (52 MB)2019-04-1890019278892 (6 MB)2019-04-18802413292 (19 MB)2019-04-1781016291233 (26 MB)2019-04-1681016291233 (24 MB)2019-04-1680016080533 (24 MB)2019-04-1681016289433 (4 MB)2019-04-08802415666 (72 MB)2019-04-089001922,76466 (5 MB)2019-04-08802415066
Dee Dee Luu et. al_The immunogenic microbial peptide RaxX represents an unclassified group of sulfated ribosomally synthesized and post-translationally modified (1 MB)2019-04-062021318 (75 MB)2019-04-029001922,76466 (2 MB)2019-04-01802417436 (39 MB)2019-04-019001922,76236 (5 MB)2019-03-28802416266 (6 MB)2019-03-27802417182 (98 MB)2019-03-279001922,76284 (7 MB)2019-03-13802416596
HLF with (176 KB)2019-03-0864345451
U87MG with (429 KB)2019-03-0891131181181
IgA with (358 KB)2019-03-0881231251251
HNE with (174 KB)2019-03-0863638381
PGM OG with (370 KB)2019-03-0821041061061
IgG with (228 KB)2019-03-0875961611
Fet NG with (229 KB)2019-03-0876063631
CBHI OG with (93 KB)2019-03-0821820201
CBHI NG with (109 KB)2019-03-0832628281
BLF with (302 KB)2019-03-0888385851 (429 KB)2019-03-0891141191191 (40 KB)2019-03-08133181 (797 KB)2019-03-08111151510 (63 MB)2019-03-08811511634212 (6 MB)2019-03-08218183216
PRM-Identifying the N-terminal amino acid for a TAG start (3 MB)2019-03-05206583
PRM-Identifying the N-terminal amino acid for an ATG start (1 MB)2019-03-05206563 (5 MB)2019-03-015502582,2633 (5 MB)2019-03-016302602,0613 (51 MB)2019-02-26747044,411576,8660 (17 MB)2019-02-26172014,598189,6470 (27 MB)2019-02-26376023,172301,0320 (17 MB)2019-02-26172014,598189,6470 (23 MB)2019-02-041902414469 (23 MB)2019-02-031902414469 (1 GB)2019-01-309,047033,292348,20120 (118 MB)2019-01-30180511728 (244 MB)2019-01-301704713795 (2 MB)2019-01-2210107062 (2 MB)2019-01-2210107058 (2 MB)2019-01-1810107058 (2 MB)2019-01-1810107062 (815 KB)2019-01-189701949311 (815 KB)2019-01-189701949311 (758 KB)2019-01-16210422021
20160309_MouseV2B12M6& (15 MB)2019-01-1646010239039 (13 MB)2019-01-1653010851045 (294 KB)2019-01-16170341661 (334 KB)2019-01-164901004831 (32 MB)2019-01-1697019496345 (62 MB)2019-01-1616603401,69045 (17 MB)2019-01-1667013462845 (1 MB)2019-01-1616603401,6501 (19 MB)2019-01-1671014269845 (929 KB)2019-01-1616303201,5481 (305 KB)2019-01-16390924561 (31 MB)2019-01-1613102661,26045 (34 MB)2019-01-1615903201,58445 (71 MB)2019-01-1611302281,06239
20190110 - Ajinomoto (23 MB)2019-01-116034161238 (451 KB)2019-01-1080381369
20180406_180307CN1_Heavy& (2 MB)2019-01-10804214427
20180401_180307CN1_Heavy& (1 MB)2019-01-10804215027
20180329_180307CN1_Heavy& (558 KB)2019-01-1080421569