期刊
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
卷 1820, 期 9, 页码 1420-1428出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagen.2011.07.004
关键词
Glycoproteins; Mass spectrometry; O-glycans; Oligosaccharides
资金
- Netherlands Genomics Initiative [93518016]
Background: Analysis of protein glycosylation is an important first step towards establishing the functions of glycans in health and disease. In contrast to N-glycans which are generally enzymatically released for analysis, there is no corresponding enzyme for O-glycan liberation. Therefore, O-glycans are generally released by chemical methods involving tedious procedures. Methods: Here, a straightforward method for the combined release and labeling of O-linked glycans from glycoproteins is described. Dimethylamine serves as the releasing agent, and 1-phenyl-3-methyl-5-pyrazolone (PMP) is employed for a prompt reaction with the reducing end of the freshly released O-glycan structures via an aldol condensation followed by a Michael-type addition resulting in a 2:1 stoichiometry of PMP per glycan. Samples are analyzed by nanoLC coupled to mass spectrometry. Results: Mucin from bovine submaxillary gland was used as a model protein to evaluate and optimize the approach that was further applied to bile salt stimulated lipase (BSSL) isolated from human milk Next to previously reported O-glycan structures two additional oligosaccharides could be detected for BSSL General Significance: In conclusion, the facile protocol established is suitable for the analysis of complex O-linked oligosaccharides from various biological samples. This article is part of a Special Issue entitled Glycoproteomics. (C) 2011 Elsevier B.V. All rights reserved.
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