Evaluation of Different N-Glycopeptide Enrichment Methods for N-Glycosylation Sites Mapping in Mouse Brain.
N-Glycosylation of proteins plays a critical role in many biological pathways. Because highly heterogeneous N-glycopeptides are present in biological sources, the enrichment procedure is a crucial step for mass spectrometry analysis. Five enrichment methods, including IP-ZIC-HILIC, hydrazide chemistry, lectin affinity, ZIC-HILIC-FA, and TiO2 affinity were evaluated and compared in the study of mapping N-glycosylation sites in mouse brain. On the basis of our results, the identified N-glycosylation sites were 1891, 1241, 891, 869, and 710 and the FDR values were 3.29, 5.62, 9.54, 9.54, and 20.02%, respectively. Therefore, IP-ZIC-HILIC enrichment method displayed the highest sensitivity and specificity. In this work, we identified a total of 3446 unique glycosylation sites conforming to the N-glycosylation consensus motif (N-X-T/S/C; X ≠ P) with (18)O labeling in 1597 N-glycoproteins. N-glycosylation site information was used to confirm or correct the transmembrane topology of the 57 novel transmembrane N-glycoproteins.
['Zhang|Chengqian|C|', 'Ye|Zilu|Z|', 'Xue|Peng|P|', 'Shu|Qingbo|Q|', 'Zhou|Yue|Y|', 'Ji|Yanlong|Y|', 'Fu|Ying|Y|', 'Wang|Jifeng|J|', 'Yang|Fuquan|F|']
D020816:Amino Acid Motifs D000818:Animals D001665:Binding Sites D001921:Brain D006020:Glycopeptides D006031:Glycosylation D007553:Isotope Labeling D013058:Mass Spectrometry D051379:Mice D010103:Oxygen Isotopes D012680:Sensitivity and Specificity
N-glycosylation sites mouse brain hydrazide lectins TiO2 ZIC-HILIC