Real-time metabolome profiling of the metabolic switch between starvation and growth
Real-time metabolome profiling of the metabolic switch between starvation and growth
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DOI：
10.1038/nmeth.3584
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发表时间：
2015-11-01
期刊：
影响因子：
48
通讯作者：
Sauer, Uwe
中科院分区：
文献类型：
--
作者：
Link, Hannes;Fuhrer, Tobias;Sauer, Uwe
代谢系统通常是对环境变化做出反应的第一个网络，监测代谢物动力学的能力是理解这些细胞反应的关键。由于监测代谢组变化在实验上繁琐且要求严格，因此从几秒到几小时的时间尺度上的动态数据很少。在这里，我们描述了通过将活细菌，酵母或哺乳动物细胞直接注射到高分辨率质谱仪中来进行实时代谢组分析，该质谱仪可以在几个小时内以15-30秒的周期自动监测约300种化合物。我们观察到能量昂贵的生物质代谢产物在大肠杆菌中的碳饥饿诱导的稳定期的积累，以及快速使用这些代谢产物后，生长恢复。通过结合实时代谢组分析与建模和抑制剂实验，我们获得的证据开关样反馈抑制氨基酸生物合成和控制底物的可用性，通过优先使用代谢上更便宜的一步补救途径，而不是昂贵的十步从头嘌呤生物合成过程中的生长恢复。
Metabolic systems are often the first networks to respond to environmental changes, and the ability to monitor metabolite dynamics is key for understanding these cellular responses. Because monitoring metabolome changes is experimentally tedious and demanding, dynamic data on time scales from seconds to hours are scarce. Here we describe real-time metabolome profiling by direct injection of living bacteria, yeast or mammalian cells into a high-resolution mass spectrometer, which enables automated monitoring of about 300 compounds in 15-30-s cycles over several hours. We observed accumulation of energetically costly biomass metabolites in Escherichia coli in carbon starvation induced stationary phase, as well as the rapid use of these metabolites upon growth resumption. By combining real-time metabolome profiling with modeling and inhibitor experiments, we obtained evidence for switch-like feedback inhibition in amino acid biosynthesis and for control of substrate availability through the preferential use of the metabolically cheaper one-step salvaging pathway over costly ten-step de novo purine biosynthesis during growth resumption.