O- and N-glycosylation analysis of cell lines by ultrahigh resolution MALDI-FTICR-MS

Vreeker, G.C.M.; Nicolardi, S.; Madunic, K.; Kotsias, M.; Burgt, Y.E.M. van der; Wuhrer, M.

doi:doi:10.1016/j.ijms.2019.116267

Glycosylation analysis from biological samples is often challenging due to the high complexity of the glycan structures found in these samples. In the present study N- and O- glycans from human colorectal cancer cell lines and human plasma were analyzed using ultrahigh resolution MALDI-FTICR-MS. N-glycans were enzymatically released from cell lines and plasma proteins, whereas beta-elimination was used for the release of O-glycans from the cells. The purified samples were mass analyzed using a 15T MALDI-FTICR-MS system, with additional MS/MS (collision-induced dissociation) experiments for O-glycan identifications. A total of 104 O-glycan and 62 N-glycan compositions were observed in the spectra obtained from colorectal cancer cell line samples. In the cell line N-glycan spectra, the highest intensity signals originated from high-mannose glycans, next to the presence of various complex type glycans. Notably, in the O-glycan spectra mono- and disaccharide signals were observed, which...Show moreGlycosylation analysis from biological samples is often challenging due to the high complexity of the glycan structures found in these samples. In the present study N- and O- glycans from human colorectal cancer cell lines and human plasma were analyzed using ultrahigh resolution MALDI-FTICR-MS. N-glycans were enzymatically released from cell lines and plasma proteins, whereas beta-elimination was used for the release of O-glycans from the cells. The purified samples were mass analyzed using a 15T MALDI-FTICR-MS system, with additional MS/MS (collision-induced dissociation) experiments for O-glycan identifications. A total of 104 O-glycan and 62 N-glycan compositions were observed in the spectra obtained from colorectal cancer cell line samples. In the cell line N-glycan spectra, the highest intensity signals originated from high-mannose glycans, next to the presence of various complex type glycans. Notably, in the O-glycan spectra mono- and disaccharide signals were observed, which are difficult to detect using alternative glycomic platforms such as porous graphitized carbon LC-MS. In the N-glycan spectra from plasma, isobaric species were resolved in MALDI-FTICR-MS spectra using absorption mode whereas these overlapped in magnitude mode. The use of ultrahigh resolution MALDI-FTICR-MS for the analysis of glycans in complex mixtures enables us to confidently analyze glycans in the matrix region of the spectrum and to differentiate isobaric glycan species. (C) 2019 The Authors. Published by Elsevier B.V.Show less

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O- and N-glycosylation analysis of cell lines by ultrahigh resolution MALDI-FTICR-MS