Integrating MS1 and MS2 Scans in High-Resolution Parallel Reaction Monitoring Assays for Targeted Metabolite Quantification and Dynamic 13C-Labeling Metabolism Analysis
Integrating MS1 and MS2 Scans in High-Resolution Parallel Reaction Monitoring Assays for Targeted Metabolite Quantification and Dynamic 13C-Labeling Metabolism Analysis
复制标题
将 MS1 和 MS2 扫描集成到高分辨率平行反应监测分析中，以进行目标代谢物定量和动态 13C 标记代谢分析。
DOI：
10.1021/acs.analchem.6b03947
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发表时间：
2017-01-03
影响因子：
7.4
通讯作者：
Shui, Wenqing
中科院分区：
文献类型：
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作者：
Li, Zhucui;Li, Yujing;Shui, Wenqing
对复杂生物材料中的目标代谢物，特别是微量代谢物和结构异构体进行定量是代谢组学面临的持续挑战。最初是为蛋白质组分析而开发的平行反应监测(PRM)技术利用了高分辨率的MS2片段离子数据，在靶向代谢物定量方面显示出很高的前景。值得注意的是，作为每个PRM扫描周期的一部分独立获取的MS1离子强度数据在PRM分析中通常未得到充分利用。在这项研究中，我们开发了一种MS1/MS2结合的PRM工作流程，用于在正交QqTOF系统上对中枢碳代谢中间体、氨基酸和莽草酸途径相关代谢物进行定量。浓度曲线评估显示，在PRM分析中同时使用MS1和MS2扫描比依赖任何一种扫描模式进行定量具有更高的灵敏度、更宽的动态范围和更好的重复性。此外，Skyline还被加入到我们的工作流程中来处理MS1/MS2离子强度数据，并消除噪声信号和带有干扰的跃迁。这种整合的MS1/MS2 PRM方法被应用于工程菌中目标代谢物的定量，以了解代谢途径的调控。此外，这种新的方法，当第一次在动态13C标记实验中实施时，在捕捉和校正同位素标记曲线以促进非静态13C标记代谢分析方面显示了其独特的优势。
Quantification of targeted metabolites, especially trace metabolites and structural isomers, in complex biological materials is an ongoing challenge for metabolomics. Initially developed for proteomic analysis, the parallel reaction monitoring (PRM) technique exploiting high-resolution MS2 fragment ion data has shown high promise for targeted metabolite quantification. Notably, MS1 ion intensity data acquired independently as part of each PRM scan cycle are often underutilized in the PRM assay. In this study, we developed an MS1/MS2-combined PRM workflow for quantification of central carbon metabolism intermediates, amino acids and shikimate pathway-related metabolites on an orthogonal QqTOF system. Concentration curve assessment revealed that exploiting both MS1 and MS2 scans in PRM analysis afforded higher sensitivity, wider dynamic range and better reproducibility than relying on either scan mode for quantification. Furthermore, Skyline was incorporated into our workflow to process the MS1/MS2 ion intensity data, and eliminate noisy signals and transitions with interferences. This integrated MS1/MS2 PRM approach was applied to targeted metabolite quantification in engineered E. coli strains for understanding of metabolic pathway modulation. In addition, this new approach, when first implemented in a dynamic 13C-labeling experiment, showed its unique advantage in capturing and correcting isotopomer labeling curves to facilitate nonstationary 13C-labeling metabolism analysis.