科研之友
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摘要
Post-translational modifications (PTMs) of serine and threonine occur by diverse mechanisms, including phosphorylation, sulfation, and various types of sugar chain modifications, making characterization of the resulting structures very labor-intensive. Moreover, to fully understand the biological functions of PTMs, both the sites of modification and the modified structures must be analyzed. The present work describes a novel, versatile strategy in which the released O-glycan and the formerly glycosylated/phosphorylated peptide are labeled and thus amenable to further study. In this approach, glycopeptides/phosphopeptides are subjected to beta-elimination in the presence of pyrazolone derivatives (BEP), which in the same reaction labels the formerly glycosylated/phosphorylated peptide. The reaction is essentially a beta-elimination/Michael addition in which a carbon-carbon bond-forming Michael donor rather than a heteroatomic Michael donor is used. The O-glycans released upon BEP are recovered as bis-pyrazolone derivatives, without any detectable side reaction (peeling). Using this technique, the O-glycan profiles of model mucin-type glycoproteins were successfully analyzed. The BEP strategy discriminates between phosphorylated and GIcNAcylated peptides, since cleaved GIcNAc is detectable. In addition, both the released O-glycan and the formerly glycosylated peptide can be selectively labeled by different reagents via a beta-elimination reaction performed in the presence of pyrazolone and the thiol Michael donor.
- 出版日期2011-12-1
