Fujita Koki, Bunyu Yasufumi, Kuroda Ken'ich, Ashitani Tatsuya, Shigeto Jun, Tsutsumi Yuji
Department of Agro-Environmental Sciences, Faculty of Agriculture, Graduate School of Kyushu University, Hakozaki 6-10-1, Higashi-ku, Fukuoka 812-8581, Japan.
Department of Agro-Environmental Sciences, Faculty of Agriculture, Graduate School of Kyushu University, Hakozaki 6-10-1, Higashi-ku, Fukuoka 812-8581, Japan.
J Plant Physiol. 2014 May 1;171(8):610-4. doi: 10.1016/j.jplph.2013.12.016. Epub 2014 Mar 20.
β-Thujaplicin is a wood monoterpene and tropolone compound with a unique conjugated 7-membered ring. Because of its strong antifungal and antitumor activities, β-thujaplicin is used in several fields. The biosynthesis pathway of β-thujaplicin has not yet been elucidated. Using Cupressus lusitanica cell cultures in a radioisotope feeding experiment, our group previously demonstrated that geranyl pyrophosphate (GPP) is the starting material of β-thujaplicin biosynthesis. The results of our previous terpene synthase assay suggested that terpinolene is the first olefin terpenoid intermediate from GPP to β-thujaplicin, although there was no experimental evidence of this at that time. In the present study, we fed deuterium-labeled terpinolene to cultured C. lusitanica cells to determine whether terpinolene is an intermediate metabolite of β-thujaplicin biosynthesis. A gas chromatography-mass spectroscopy analysis of the cell extracts from labeled terpinolene cultures revealed a peak of labeled β-thujaplicin that was not observed after treatment with non-labeled terpinolene. The identification of labeled β-thujaplicin was also performed by mass spectrum assignment. The outcome indicated that terpinolene is indeed an intermediate metabolite of β-thujaplicin biosynthesis. To the best of our knowledge, there has been no prior report that tropolone compounds are biosynthesized via a terpene biosynthesis system, and our results thus suggest the existence of a novel biosynthetic pathway that produces the conjugated 7-membered ring.
β-崖柏素是一种具有独特共轭七元环的木本单萜和托酚酮化合物。由于其强大的抗真菌和抗肿瘤活性，β-崖柏素被应用于多个领域。β-崖柏素的生物合成途径尚未阐明。在一项放射性同位素饲喂实验中，我们的研究小组利用葡萄牙柏细胞培养物，先前已证明香叶基焦磷酸（GPP）是β-崖柏素生物合成的起始原料。我们之前的萜类合酶测定结果表明，萜品油烯是从GPP到β-崖柏素的首个烯烃类萜类中间体，尽管当时尚无这方面的实验证据。在本研究中，我们将氘标记的萜品油烯饲喂给培养的葡萄牙柏细胞，以确定萜品油烯是否为β-崖柏素生物合成的中间代谢物。对来自标记萜品油烯培养物的细胞提取物进行气相色谱-质谱分析，结果显示出一个标记β-崖柏素的峰，在用未标记的萜品油烯处理后未观察到该峰。标记β-崖柏素的鉴定也通过质谱归属进行。结果表明，萜品油烯确实是β-崖柏素生物合成的中间代谢物。据我们所知，此前尚无关于托酚酮化合物通过萜类生物合成系统进行生物合成的报道，因此我们的结果表明存在一条产生共轭七元环的新型生物合成途径。
