Cleavable hydrophobic derivatization strategy for enrichment and identification of phosphorylated lysine peptides
Cleavable hydrophobic derivatization strategy for enrichment and identification of phosphorylated lysine peptides
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DOI：
10.1007/s00216-019-01770-w
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发表时间：
2019-07-01
影响因子：
4.3
通讯作者：
Zhang, Yukui
中科院分区：
文献类型：
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作者：
Hu, Yechen;Li, Yang;Zhang, Yukui
由于结构的灵活性和对酸的不稳定性，磷酸化赖氨酸(PLYS)的鉴定是一个巨大的挑战。我们在这里报告了一种可切割疏水衍生化(CHD)策略，用于浓缩和鉴定PLYS多肽。首先合成2，5-dioxopyrrolidin-1-yl-3-(decyldisulfanyl)propanoate与去磷酸化的赖氨酸多肽反应，然后用C-18柱捕获衍生多肽，然后切割疏水链，并在目标多肽上留下特定的标记以供进一步鉴定。通过CHD，实现了从1：1000质量比的干扰肽中富集PLYS多肽。此外，利用CHD从大肠杆菌裂解物中筛选PLYS靶点，从35个蛋白质中鉴定出39个PLYS位点。基因本体论(GO)分析表明，这些蛋白质在分解代谢、代谢、生物发生和生物合成过程中发挥着重要作用。所有这些结果表明，CHD可能为全面分析PLYS蛋白质组奠定了基础。
Because of structural flexibility and acid lability, the identification of phosphorylated lysine (pLys) peptides is a great challenge. We report here a cleavable hydrophobic derivatization (CHD) strategy for the enrichment and identification of pLys peptides. First, 2,5-dioxopyrrolidin-1-yl-3-(decyldisulfanyl)propanoate was synthesized to react with dephosphorylated lysine peptides, and then the derived peptides were captured by a C-18 column, followed by cleavage of the hydrophobic chain, with the specific label left on the target peptides for further identification. By CHD, the enrichment of pLys peptides from interfering peptides (1:1000 mass ratio) was achieved. Furthermore, CHD was applied to screen the pLys targets from Escherichia coli lysates, and 39 pLys sites from 35 proteins were identified. Gene Ontology (GO) analysis showed that these proteins played vital roles in catabolism, metabolism, biogenesis, and biosynthetic processes. All these results demonstrate that CHD might pave the way for comprehensive profiling of the pLys proteome.