An immobilized titanium (IV) ion affinity chromatography adsorbent for solid phase extraction of phosphopeptides for phosphoproteome analysis
An immobilized titanium (IV) ion affinity chromatography adsorbent for solid phase extraction of phosphopeptides for phosphoproteome analysis
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用于固相萃取磷酸肽进行磷酸蛋白质组分析的固定化钛 (IV) 离子亲和色谱吸附剂
DOI：
10.1016/j.chroma.2017.03.026
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发表时间：
2017
影响因子：
4.1
通讯作者：
Zou Hanfa
中科院分区：
文献类型：
--
作者：
Yao Yating;Dong Jing;Dong Mingming;Liu Fangjie;Wang Yan;Mao Jiawei;Ye Mingliang;Zou Hanfa
在这项研究中，我们开发了一种离心辅助固相萃取（SPE）的方法选择性富集磷酸肽使用一种新的Ti4+-IMAC材料在内部合成。该材料具有粒径大、比表面积大的特点，更适合于固相萃取富集磷酸肽。应用负载有Ti 4 +-IMAC材料的自旋尖端从复杂蛋白质酶中富集磷酸肽。结果表明，牛血清白蛋白和β-酪蛋白的胰蛋白酶酶解物可以特异性地富集磷酸肽，其摩尔比可达1000：1。从HeLa细胞蛋白的胰蛋白酶消化物中可以鉴定出约4700个独特的磷酸化肽，特异性高达99%。该吸头被证明具有良好的柱间重现性。此外，它适合于分析微量样品。与传统的溶液法相比，固相萃取法能快速、完全地将物质与溶液分离，是一种省时、方便的磷酸肽富集方法。与商业TiO 2材料相比，该材料具有更高的磷酸肽识别率和更高的富集特异性。
In this study, we developed a centrifugation assisted solid phase extraction (SPE) method for the selective enrichment of phosphopeptides using a new Ti4+-IMAC material synthesized in-house. This new material has the feature of big size and large specific surface area which makes it more suitable to enrich phosphopeptides in a SPE way. The spin tips loaded with the Ti4+-IMAC material were applied to enrich phosphopeptides from the complex protein digests. It was found that phosphopeptides can be specifically enriched from tryptic digest of bovine serum albumin and β-casein at a molar ratio up to 1000:1. And about 4700 unique phosphorylated peptides can be identified with the specificity as high as 99% from the tryptic digest of HeLa cell proteins. This tip was demonstrated to have good column-to-column reproducibility. Furthermore, it is fitted to analyze minute amount of sample. Compared with the conventional solution method, the SPE method facilitated the rapid and complete separation of the material with solution, which making it a time-saving and convenient method for phosphopeptide enrichment. Compared with the commercial TiO2material, this new materials yielded much more phosphopeptide identifications and much higher enrichment specificity.