ABSTRACT The gut microbiota synthesize hundreds of molecules, many of which are known to impact host physiology. Among the most abundant metabolites are the secondary bile acids deoxycholic acid (DCA) and lithocholic acid (LCA), which accumulate at ~500 μM and are known to block C. difficile growth 1 , promote hepatocellular carcinoma 2 , and modulate host metabolism via the GPCR TGR5 3 . More broadly, DCA, LCA and their derivatives are a major component of the recirculating bile acid pool 4 ; the size and composition of this pool are a target of therapies for primary biliary cholangitis and nonalcoholic steatohepatitis. Despite the clear impact of DCA and LCA on host physiology, incomplete knowledge of their biosynthetic genes and a lack of genetic tools in their native producer limit our ability to modulate secondary bile acid levels in the host. Here, we complete the pathway to DCA/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 8-step conversion of cholic acid to DCA. We then engineer the pathway into Clostridium sporogenes , conferring production of DCA and LCA on a non-producing 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, and provide a road map for deorphaning and engineering pathways from the microbiome as a critical step toward controlling the metabolic output of the gut microbiota.

Consuming diets rich in plant versus animal products changes the microbes found in the human gut within days, with important implications for our health and evolution. Diet influences the structure and function of the gut microbiota in the long term, but it is not clear how rapidly the microbiota is affected by short-term dietary change. Peter Turnbaugh and colleagues studied the effect of transition to a diet consisting entirely of either animal products or plant products on the composition and function of the human gut microbiota. They find that the community changes rapidly, within a single day, overwhelming the pre-existing inter-individual differences in microbiota composition to recapitulate expected patterns of composition and metabolic function for carnivorous and herbivorous mammals. The animal-based diet was associated with higher levels of bile-tolerant microorganisms, including the bacterium Bilophila wadsworthia, which has previously been linked to inflammatory bowel disease. The authors also detected intact foodborne fungi, bacteria and viruses in the distal gut. Long-term dietary intake influences the structure and activity of the trillions of microorganisms residing in the human gut1,2,3,4,5, but it remains unclear how rapidly and reproducibly the human gut microbiome responds to short-term macronutrient change. Here we show that the short-term consumption of diets composed entirely of animal or plant products alters microbial community structure and overwhelms inter-individual differences in microbial gene expression. The animal-based diet increased the abundance of bile-tolerant microorganisms (Alistipes, Bilophila and Bacteroides) and decreased the levels of Firmicutes that metabolize dietary plant polysaccharides (Roseburia, Eubacterium rectale and Ruminococcus bromii). Microbial activity mirrored differences between herbivorous and carnivorous mammals2, reflecting trade-offs between carbohydrate and protein fermentation. Foodborne microbes from both diets transiently colonized the gut, including bacteria, fungi and even viruses. Finally, increases in the abundance and activity of Bilophila wadsworthia on the animal-based diet support a link between dietary fat, bile acids and the outgrowth of microorganisms capable of triggering inflammatory bowel disease6. In concert, these results demonstrate that the gut microbiome can rapidly respond to altered diet, potentially facilitating the diversity of human dietary lifestyles.