O-glycosylation sites identified from mucin core-1 type glycopeptides from human serum.
O-glycosylation sites identified from mucin core-1 type glycopeptides from human serum.
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DOI：
10.1007/s10719-015-9630-6
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发表时间：
2016-06
影响因子：
3
通讯作者：
Medzihradszky KF
中科院分区：
文献类型：
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作者：
Darula Z;Sarnyai F;Medzihradszky KF
在这项工作中，从人血清中富集了具有粘蛋白核心-1型结构的O-连接糖肽。由于人血清中约70%的O-聚糖与植物凝集素Jacalin结合，我们测试了先前成功的方案，该方案将Jacalin在蛋白质和肽水平上的亲和富集与ERLIC色谱结合作为中间的进一步富集步骤，以消除未修饰肽的高背景。同时，我们开发了一种更简单，更快的新工作流程，依次使用两种凝集素：小麦胚芽凝集素和Jacalin。第一种凝集素提供一般糖肽富集，而第二种特异性富集具有Galβ1-3GalNAcα结构的O-连接糖肽。富集样品的质谱分析表明，新的样品制备方法比前者具有更高的选择性和灵敏度。总共，在这项研究中，在20个蛋白质中鉴定了52个独特的糖基化位点。
In this work O-linked glycopeptides bearing mucin core-1 type structures were enriched from human serum. Since about 70% of the O-glycans in human serum bind to the plant lectin Jacalin, we tested a previously successful protocol that combined Jacalin affinity enrichment on the protein- and peptide-level with ERLIC chromatography as a further enrichment step in between, to eliminate the high background of unmodified peptides. In parallel, we developed a simpler and significantly faster new workflow that used two lectins sequentially: wheat germ agglutinin and then Jacalin. The first lectin provides general glycopeptide enrichment, while the second specifically enriches O-linked glycopeptides with Galβ1–3GalNAcα structures. Mass spectrometric analysis of enriched samples showed that the new sample preparation method is more selective and sensitive than the former. Altogether, 52 unique glycosylation sites in 20 proteins were identified in this study.