Lampe Sarah, Fiedler Dorothea
Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany.
Humboldt-Universität zu Berlin, Institut für Chemie, Berlin, Germany.
Methods Mol Biol. 2025;2972:135-152. doi: 10.1007/978-1-0716-4799-8_11.
The proteome-wide detection of different posttranslational modifications (PTMs) still poses a notable analytical challenge. Nevertheless, the identification of protein pyrophosphorylation sites using mass spectrometry (MS) was reported recently. The implementation of an enrichment workflow proved to be key to characterize this substoichiometric modification within complex cell lysate samples. Analysis of the enriched samples using data-dependent neutral-loss triggered electron transfer/higher-energy collision dissociation (DDNL EThcD) MS enabled the detection of endogenous protein pyrophosphorylation. In this chapter, the bottom-up pyrophosphoproteomics analysis, including an enrichment workflow, to identify protein pyrophosphorylation sites is described.
对不同翻译后修饰（PTM）进行全蛋白质组检测仍然是一项重大的分析挑战。然而，最近有报道称利用质谱（MS）鉴定蛋白质焦磷酸化位点。富集工作流程的实施被证明是在复杂细胞裂解物样品中表征这种亚化学计量修饰的关键。使用数据依赖型中性丢失触发电子转移/高能碰撞解离（DDNL EThcD）质谱对富集样品进行分析，能够检测内源性蛋白质焦磷酸化。在本章中，将描述用于鉴定蛋白质焦磷酸化位点的自下而上的焦磷酸蛋白质组学分析，包括富集工作流程。
