Sequential enrichment of singly- and multiply-phosphorylated peptides with zwitterionic hydrophilic interaction chromatography material.
Sequential enrichment of singly- and multiply-phosphorylated peptides with zwitterionic hydrophilic interaction chromatography material.
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DOI：
10.1016/j.chroma.2015.08.018
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发表时间：
2015-09
期刊：
影响因子：
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通讯作者：
Qianying Sheng;Kaiya Yang;X. Xue;Xiuling Li;Zhimou Guo;Aijin Shen;Yanxiong Ke;M. Lan;Xinmiao Liang
中科院分区：
文献类型：
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作者：
Qianying Sheng;Kaiya Yang;X. Xue;Xiuling Li;Zhimou Guo;Aijin Shen;Yanxiong Ke;M. Lan;Xinmiao Liang
提出了一种有趣的、新颖的方法，用一种两性离子材料“Click TE-Cys”来选择性地和顺序地富集单和多个磷酸化肽。通过考察乙腈含量、pH值和缓冲液浓度对磷酸肽保留的影响，系统地研究了磷酸肽与Click TE-Cys之间的保留机理。亲水相互作用和静电相互作用都参与了磷酸肽与Click TE-Cys之间的相互作用。在此基础上，建立了优化的Click TE-Cys选择性富集磷小肽的方法。该方法不仅对磷酸多肽具有很高的选择性，而且可以将单磷酸化肽和多磷酸化肽分成两个组分。该方法使用相对复杂的样品进行评估，包括摩尔比为1：10的α-酪蛋白和牛血清白蛋白的多肽混合物和脱脂牛奶。这种高效、优化的方法具有丰富多聚磷酸化多肽的潜力，是磷酸蛋白质组分析中特异性富集磷酸多肽的有价值的工具。
An interesting and novel method for the selective and sequential enrichment of singly- and multiply-phosphorylated peptides with a zwitterionic material “Click TE-Cys” is presented. Retention mechanisms between phosphopeptides and Click TE-Cys are systematically investigated by checking the influence of acetonitrile content, pH value, and buffer concentration on the retention of phosphopeptides. Both hydrophilic interaction and electrostatic interaction are involved in retention between phosphopeptides and Click TE-Cys. Based on these results, an optimized method is established for selective enrichment of phosphopeptides using Click TE-Cys. This method not only exhibits high selectivity for phosphopeptides, but also fractionates singly- and multiply-phosphorylated peptides into two fractions. This method was evaluated using relatively complex samples, including peptide mixtures of α-casein and bovine serum albumin (BSA) at a molar ratio of 1:10 and skim milk. This efficient and optimized protocol has great potential for enriching multiply-phosphorylated peptides and could be a valuable tool for specific enrichment of phosphopeptides in phosphoproteome analysis.