The gut microbiota synthesize hundreds of molecules, many of which influence host physiology. Among the most abundant metabolites are the secondary bile acids deoxycholic acid (DCA) and lithocholic acid (LCA), which accumulate at concentrations of around 500 μM and are known to block the growth of Clostridium difficile 1 , promote hepatocellular carcinoma 2 and modulate host metabolism via the G-protein-coupled receptor TGR5 (ref. 3 ). More broadly, DCA, LCA and their derivatives are major components of the recirculating pool of bile acids 4 ; the size and composition of this pool are a target of therapies for primary biliary cholangitis and nonalcoholic steatohepatitis. Nonetheless, despite the clear impact of DCA and LCA on host physiology, an incomplete knowledge of their biosynthetic genes and a lack of genetic tools to enable modification of their native microbial producers limit our ability to modulate secondary bile acid levels in the host. Here we complete the pathway to DCA and LCA by assigning and characterizing enzymes for each of the steps in its reductive arm, revealing a strategy in which the A–B rings of the steroid core are transiently converted into an electron acceptor for two reductive steps carried out by Fe–S flavoenzymes. Using anaerobic in vitro reconstitution, we establish that a set of six enzymes is necessary and sufficient for the eight-step conversion of cholic acid to DCA. We then engineer the pathway into Clostridium sporogenes , conferring production of DCA and LCA on a nonproducing commensal and demonstrating that a microbiome-derived pathway can be expressed and controlled heterologously. These data establish a complete pathway to two central components of the bile acid pool. The biosynthetic pathway that produces the secondary bile acids DCA and LCA in human gut microbes has been fully characterized, engineered into another bacterial host, and used to confer DCA production in germ-free mice—an important proof-of-principle for the engineering of gut microbial pathways.

中文翻译：

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肠道微生物组胆汁酸脱羟基的代谢途径

肠道微生物群合成数百种分子，其中许多影响宿主生理。最丰富的代谢物包括次级胆汁酸脱氧胆酸 (DCA) 和石胆酸 (LCA)，它们以约 500 μM 的浓度累积，已知可阻断艰难梭菌 1 的生长，促进肝细胞癌 2 和调节宿主代谢通过 G 蛋白偶联受体 TGR5（参考文献 3）。更广泛地说，DCA、LCA 及其衍生物是胆汁酸再循环库的主要成分 4 ；该池的大小和组成是原发性胆汁性胆管炎和非酒精性脂肪性肝炎治疗的目标。尽管如此，尽管 DCA 和 LCA 对宿主生理有明显影响，对它们的生物合成基因的不完全了解以及缺乏能够修改其本地微生物生产者的遗传工具限制了我们调节宿主中次级胆汁酸水平的能力。在这里，我们通过为还原臂中的每个步骤分配和表征酶来完成 DCA 和 LCA 的途径，揭示了一种策略，其中类固醇核心的 A-B 环瞬时转化为电子受体，用于携带的两个还原步骤由 Fe-S 黄素酶产生。使用厌氧体外重组，我们确定一组六种酶对于胆酸向 DCA 的八步转化是必要且充分的。然后我们将途径设计成产孢梭菌，将 DCA 和 LCA 的生产赋予非生产性共生体，并证明微生物组衍生途径可以异源表达和控制。这些数据建立了通往胆汁酸池两个中心成分的完整途径。在人类肠道微生物中产生次级胆汁酸 DCA 和 LCA 的生物合成途径已得到充分表征，被工程化到另一个细菌宿主中，并用于在无菌小鼠中产生 DCA——这是工程化的重要原理证明肠道微生物通路。