Miller Garret P, Mutabdžija-Nedelcheva Lana, Andersen Trine B, Pascoe Imani, Van Winkle Kathryn, Pluskal Tomáš, Hamberger Björn
Biochemistry and Molecular Biology, Michigan State University, East Lansing, Michigan, United States.
Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo nám. 2, 160 00 Prague, Czechia.
bioRxiv. 2025 May 17:2025.05.15.654307. doi: 10.1101/2025.05.15.654307.
Roots from the (Wolf's-Bane) and (Larkspur) genera have been widely used in traditional medicine owing to the abundance of bioactive diterpenoid alkaloids that they produce. Despite extensive research on these compounds and their potential medicinal applications, their structural complexity precludes their production through total chemical synthesis, and little progress has been made towards elucidation of their biosynthetic pathways. Here, we report the entry steps in the biosynthesis of the diterpenoid alkaloid atisinium, constituting six enzymes identified from the Siberian Larkspur () and garden monkshood () through a combination of comparative transcriptomics between tissue types and genera and coexpression analysis. This pathway includes a pair of terpene synthases, three cytochromes P450, and a reductase with little homology to other characterized enzymes. We further demonstrate, through incorporation of isotopically labelled substrates, the preference of the reductase for ethanolamine over ethylamine, and similarly that ethanolamine is the preferred source of nitrogen for the majority of detected diterpenoid alkaloids. Identification of these enzymes and production of a key intermediate in a heterologous host paves the way for biosynthetic production of this group of metabolites with promise for medicinal applications.
由于含有丰富的生物活性二萜生物碱，菟葵属和翠雀属植物的根在传统医学中被广泛应用。尽管对这些化合物及其潜在药用价值进行了广泛研究，但因其结构复杂，无法通过全化学合成来制备，且在阐明其生物合成途径方面进展甚微。在此，我们报告了二萜生物碱阿替生的生物合成起始步骤，通过组织类型和属间的比较转录组学及共表达分析相结合，从西伯利亚翠雀和川乌中鉴定出了六种酶。该途径包括一对萜类合酶、三种细胞色素P450和一种与其他已鉴定酶几乎没有同源性的还原酶。我们通过掺入同位素标记底物进一步证明，该还原酶对乙醇胺的偏好高于乙胺，同样，乙醇胺是大多数检测到的二萜生物碱的首选氮源。这些酶的鉴定以及在异源宿主中产生关键中间体，为有望用于医药应用的这组代谢物的生物合成生产铺平了道路。
