Department of Chemical Engineering, Stanford University, Stanford, CA 94305.
HHMI, Stanford University, Stanford, CA 94305.
Proc Natl Acad Sci U S A. 2021 Jun 15;118(24). doi: 10.1073/pnas.2102949118.
Plants synthesize many diverse small molecules that affect function of the mammalian central nervous system, making them crucial sources of therapeutics for neurological disorders. A notable portion of neuroactive phytochemicals are lysine-derived alkaloids, but the mechanisms by which plants produce these compounds have remained largely unexplored. To better understand how plants synthesize these metabolites, we focused on biosynthesis of the Lycopodium alkaloids that are produced by club mosses, a clade of plants used traditionally as herbal medicines. Hundreds of Lycopodium alkaloids have been described, including huperzine A (HupA), an acetylcholine esterase inhibitor that has generated interest as a treatment for the symptoms of Alzheimer's disease. Through combined metabolomic profiling and transcriptomics, we have identified a developmentally controlled set of biosynthetic genes, or potential regulon, for the Lycopodium alkaloids. The discovery of this putative regulon facilitated the biosynthetic reconstitution and functional characterization of six enzymes that act in the initiation and conclusion of HupA biosynthesis. This includes a type III polyketide synthase that catalyzes a crucial imine-polyketide condensation, as well as three Fe(II)/2-oxoglutarate-dependent dioxygenase (2OGD) enzymes that catalyze transformations (pyridone ring-forming desaturation, piperidine ring cleavage, and redox-neutral isomerization) within downstream HupA biosynthesis. Our results expand the diversity of known chemical transformations catalyzed by 2OGDs and provide mechanistic insight into the function of noncanonical type III PKS enzymes that generate plant alkaloid scaffolds. These data offer insight into the chemical logic of Lys-derived alkaloid biosynthesis and demonstrate the tightly coordinated coexpression of secondary metabolic genes for the biosynthesis of medicinal alkaloids.
植物合成了许多影响哺乳动物中枢神经系统功能的不同小分子，使它们成为治疗神经疾病的重要治疗药物来源。许多具有神经活性的植物化学成分是赖氨酸衍生的生物碱，但植物产生这些化合物的机制在很大程度上仍未得到探索。为了更好地理解植物如何合成这些代谢物，我们专注于研究石松类植物产生的石松生物碱的生物合成，石松类植物是一类传统上用作草药的植物。已经描述了数百种石松生物碱，包括石杉碱 A（HupA），一种乙酰胆碱酯酶抑制剂，作为治疗阿尔茨海默病症状的药物引起了人们的兴趣。通过结合代谢组学分析和转录组学，我们已经确定了一组与发育相关的石松生物碱生物合成基因或潜在调控基因。发现这个假定的调控基因有助于对参与 HupA 生物合成起始和结束的六种酶进行生物合成重建和功能表征。这包括催化关键亚胺-聚酮缩合反应的 III 型聚酮合酶，以及催化吡啶酮环形成不饱和、哌啶环裂解和氧化还原中性异构化等转化的三种 Fe(II)/2-氧代戊二酸依赖性双加氧酶（2OGD）酶。我们的结果扩展了已知由 2OGD 催化的化学转化的多样性，并提供了对生成植物生物碱支架的非典型 III 型 PKS 酶功能的机制见解。这些数据提供了对 Lys 衍生生物碱生物合成的化学逻辑的深入了解，并证明了用于合成药用生物碱的次生代谢基因的紧密协调共表达。
