摘要
蛋白质的糖基化是最重要的翻译后修饰之一,与蛋白质结构和功能的关系密切。凝集素亲和色谱是蛋白质糖基化研究中很常用的工具,不同的凝集素可以对不同的单糖或寡糖有特异的富集作用。麦胚凝集素(W GA)由于其特异作用的糖型广泛存在而成为使用最多的凝集素之一。在本研究中,发现将W GA用于糖肽亲和富集会导致部分肽段的降解,从而导致后续的肽段序列分析的失败。本文用4种标准蛋白质对这种现象进行了验证,结果表明肽段的降解可以发生在多个位点,其中较多地发生在酪氨酸、苯丙氨酸及亮氨酸的羧基端。这一结果提示:在糖蛋白质组研究中,如果应用W GA富集糖肽并采用质谱进行鉴定,则采用半酶切或非特异性酶切的检索策略更为合适。
As one of the most important post-translational modifications （ PTMs）, glycosylation has a significant effect on the structure and functions of proteins. One of the important tools for glycoprotein research is lectins, which are known to have the ability to bind specific oligosaccharide moieties. Many different pure lectins are commercially available in an immobilized form suitable for glycoprotein purification, in which, wheat germ agglutinin （WGA） is the most popular one for its broad binding ability for different glycans. Lectins are usually used for the enrichment of glycoproteins or glycopeptides. In our research, the amide bind of peptides, including glycopeptides, can be cleavaged when the peptide mixture passes through the lectin column, which will lead a failure in sequent identification. Four standard proteins were used to verify this phenomenon and the peptide degradation was confirmed. The cleavage can occur at many positions, but more inclined to the C-terminal of the peptides with the amino acid residual of phenylalanine （Phe）, leucine （Leu） and tyrosine （Tyr）. From this result, we first revealed the problem existed in the affinity separation strategy of glycopeptides using WGA in proteomics research very popularly and suggested that using partial or no enzyme digestion database searching parameter would be more suitable for glycopeptides identification after WGA enrichment.
出处
《色谱》
CAS
CSCD
北大核心
2009年第1期19-23,共5页
Chinese Journal of Chromatography
基金
国家自然科学基金项目(20635010,20505018)
国家重点基础研究“973”计划项目(2007CB914104)
关键词
麦胚凝集素亲和色谱
糖肽
肽键
断裂
蛋白质组学
wheat germ agglutinin （ WGA ） affinity chromatography
glycopeptides
amino bond
cleavage
proteomics
