• 对细菌的7α-脱羟基代谢途径中的酶进行纯化和分析,在体外重构了该通路,解析了其中的还原反应部分;
  • 7α-脱羟基作用只需bai操纵子中的6个酶(BaiB/CD/A2/E/F/H)即可完成完整的8步反应,将胆酸转化为DCA;
  • 胆汁酸分子甾体骨架中的A-B环会被短暂转化为一种高度氧化的中间体,作为之后BaiH和BaiCD两种Fe-S黄素酶介导的关键还原反应的底物;
  • 用3个接合质粒将该通路完整转化到生孢梭菌中,获得的工程菌株具有产生DCA和LCA的能力。
Nature [IF:69.504]

A metabolic pathway for bile acid dehydroxylation by the gut microbiome



2020-06-17, Article

Abstract & Authors:展开

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 difficile1, promote hepatocellular carcinoma2 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 acids4; 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.

First Authors:
Masanori Funabashi,Tyler L Grove

Correspondence Authors:
Steven C Almo,Michael A Fischbach

All Authors:
Masanori Funabashi,Tyler L Grove,Min Wang,Yug Varma,Molly E McFadden,Laura C Brown,Chunjun Guo,Steven Higginbottom,Steven C Almo,Michael A Fischbach