Cytochrome P450 promiscuity leads to a bifurcating biosynthetic pathway for tanshinones.
Cytochrome P450 promiscuity leads to a bifurcating biosynthetic pathway for tanshinones.
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细胞色素 P450 混杂导致丹参酮生物合成途径分叉
DOI：
10.1111/nph.13790
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发表时间：
2016-04
期刊：
影响因子：
--
通讯作者：
Huang L
中科院分区：
文献类型：
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作者：
Guo J;Ma X;Cai Y;Ma Y;Zhan Z;Zhou YJ;Liu W;Guan M;Yang J;Cui G;Kang L;Yang L;Shen Y;Tang J;Lin H;Ma X;Jin B;Liu Z;Peters RJ;Zhao ZK;Huang L
细胞色素P450（CYP）在产生萜类化合物的结构多样性中起着关键作用，萜类化合物是植物天然产物中最大的一类。然而，CYP的功能表征一直是具有挑战性的，因为在植物基因组中发现的广泛的家庭，不同的反应性和难以接近的底物和产品。在这里，我们提出了两个CYP，CYP 76 AH 3和CYP 76 AK 1，这两个CYP依次作用，形成一个分叉途径的丹参酮，含氧二萜类化合物的生物合成从中国药用植物丹参（丹参）的特征。这些CYP与已知的丹参毛状根中负责丹参酮生成的基因具有相似的转录谱。生物化学和RNA干扰研究表明，这两种CYP是混杂的。CYP 76 AH 3在两个不同的碳中心氧化Ferruginol，CYP 76 AK 1羟基化两个所得中间体的C-20。这些化合物一起将ferruginol转化为11，20-二羟基ferruginol和11，20-二羟基sugiol，从而转化为丹参酮。此外，我们证明了这些CYP的工程酵母异源生产六个含氧二萜类化合物，这反过来又使CYP介导的氧化产生的三种新化合物的结构表征的效用。我们的研究结果突出了将多个CYP纳入二萜代谢工程，以及在萜类生物合成中产生复杂网络的持续趋势。
Cytochromes P450 (CYPs) play a key role in generating the structural diversity of terpenoids, the largest group of plant natural products. However, functional characterization of CYPs has been challenging because of the expansive families found in plant genomes, diverse reactivity and inaccessibility of their substrates and products. Here we present the characterization of two CYPs, CYP76AH3 and CYP76AK1, which act sequentially to form a bifurcating pathway for the biosynthesis of tanshinones, the oxygenated diterpenoids from the Chinese medicinal plant Danshen (Salvia miltiorrhiza). These CYPs had similar transcription profiles to that of the known gene responsible for tanshinone production in elicited Danshen hairy roots. Biochemical and RNA interference studies demonstrated that both CYPs are promiscuous. CYP76AH3 oxidizes ferruginol at two different carbon centers, and CYP76AK1 hydroxylates C-20 of two of the resulting intermediates. Together, these convert ferruginol into 11,20-dihydroxy ferruginol and 11,20-dihydroxy sugiol en route to tanshinones. Moreover, we demonstrated the utility of these CYPs by engineering yeast for heterologous production of six oxygenated diterpenoids, which in turn enabled structural characterization of three novel compounds produced by CYP-mediated oxidation. Our results highlight the incorporation of multiple CYPs into diterpenoid metabolic engineering, and a continuing trend of CYP promiscuity generating complex networks in terpenoid biosynthesis.