Chip-based enrichment and NanoLC-MS/MS analysis of phosphopeptides from whole lysates
Chip-based enrichment and NanoLC-MS/MS analysis of phosphopeptides from whole lysates
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DOI：
10.1021/pr700635a
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发表时间：
2008-04-01
影响因子：
4.4
通讯作者：
Heck, Albert J. R.
中科院分区：
文献类型：
--
作者：
Mohammed, Shabaz;Kraiczek, Karsten;Heck, Albert J. R.
蛋白磷酸化可能是最广泛和最重要的翻译后修饰（PTM）。考虑到这样的说法，人们已经做出了巨大的努力来改进方法，以便全面研究细胞磷酸化事件，这也就不足为奇了。然而，全面鉴定蛋白质磷酸化位点仍然是一个挑战，最好留给经验丰富的蛋白质组学专家。HPLC芯片制造的最新进展为生物分析领域的流行技术自动化创造了一个环境。一个这样的工具将受益于自动化“纳米流”分析带来的增加的易用性和信心是磷酸肽富集。为此，我们开发了一种可重复使用的高效液相色谱纳米流芯片，利用TiO2颗粒选择性富集磷酸肽。这样的设计被证明是强大的，易于使用，并且能够在包括微量复杂细胞裂解物在内的数十种分析中保持一致的性能。
Protein phosphorylation may be the most widespread and possibly most important post-translational modification (PTM). Considering such a claim, it should be no surprise that huge efforts have been made to improve methods to allow comprehensive study of cellular phosphorylation events. Nevertheless, comprehensive identification of sites of protein phosphorylation is still a challenge, best left to experienced proteomics experts. Recent advances in HPLC chip manufacturing have created an environment to allow automation of popular techniques in the bioanalytical world. One such tool that would benefit from the increased ease and confidence brought by automated 'nanoflow' analysis is phosphopeptide enrichment. To this end, we have developed a reusable HPLC nanoflow rate chip using TiO2 particles for selective phosphopeptide enrichment. Such a design proved robust, easy to use, and was capable of consistent performance over tens of analyses including minute amounts of complex cellular lysates.