Biochemical synthesis of uniformly 13C-labeled diterpene hydrocarbons and their bioconversion to diterpenoid phytoalexins in planta
Biochemical synthesis of uniformly 13C-labeled diterpene hydrocarbons and their bioconversion to diterpenoid phytoalexins in planta
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DOI：
10.1080/09168451.2017.1285689
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发表时间：
2017-02
期刊：
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通讯作者：
Zhongfeng Ye;Kazuya Nakagawa;M. Natsume;H. Nojiri;H. Kawaide;K. Okada
中科院分区：
文献类型：
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作者：
Zhongfeng Ye;Kazuya Nakagawa;M. Natsume;H. Nojiri;H. Kawaide;K. Okada
植物内酯和植物内酯是水稻体内诱导产生的主要二萜类植物防御素。尽管对这些植物保卫素的生物合成途径进行了广泛的研究，但二萜碳氢化合物的生物转化在植物中并未显示出来。为了阐明这些植物保健素的整个生物合成途径，我们用酶法合成了均一的13C标记的对位木薯二烯和合成的对位木薯二烯，并用GC-MS和13C-核磁共振对其结构进行了验证。将13C标记的底物应用于水稻叶片上，利用LC-MS/MS检测到13C标记的代谢物。进一步将该方法应用于苔藓羽状苔藓和最近的水稻种Leersia perrieri的外群，在这些植物中成功地将这些标记的底物转化为植物保卫素。这些结果表明，从这些二萜碳氢化合物到最终产物植物保健素的真正生物合成途径存在于这些植物中，并且酶促合成的[U-13C20]二萜底物是一种强大的工具，可以在不稀释天然丰富的未标记化合物的情况下追踪内源代谢物。图形摘要统一标记的13C-二萜：天然丰富的未标记化合物示踪内源性代谢物的有力工具
Phytocassanes and momilactones are the major diterpenoid phytoalexins inductively produced in rice as bioactive substances. Regardless of extensive studies on the biosynthetic pathways of these phytoalexins, bioconversion of diterpene hydrocarbons is not shown in planta. To elucidate the entire biosynthetic pathways of these phytoalexins, uniformly 13C-labeled ent-cassadiene and syn-pimaradiene were enzymatically synthesized with structural verification by GC–MS and 13C-NMR. Application of the 13C-labeled substrates on rice leaves led to the detection of 13C-labeled metabolites using LC-MS/MS. Further application of this method in the moss Hypnum plumaeforme and the nearest out-group of Oryza species Leersia perrieri, respectively, resulted in successful bioconversion of these labeled substrates into phytoalexins in these plants. These results demonstrate that genuine biosynthetic pathways from these diterpene hydrocarbons to the end product phytoalexins occur in these plants and that enzymatically synthesized [U-13C20] diterpene substrates are a powerful tool for chasing endogenous metabolites without dilution with naturally abundant unlabeled compounds. Graphical abstract Uniformly labeled 13C-diterpene: A powerful tool for tracing endogenous metabolites without dilution with naturally abundant unlabeled compounds