Metabolism of Oxo-Bile Acids and Characterization of Recombinant 12α-Hydroxysteroid Dehydrogenases from Bile Acid 7α-Dehydroxylating Human Gut Bacteria

Bile acids are important cholesterol-derived nutrient signaling hormones, synthesized in the liver, that act as detergents to solubilize dietary lipids. Bile acid 7 alpha-dehydroxylating gut bacteria generate the toxic bile acids deoxycholic acid and lithocholic acid from host bile acids. The ability of these bacteria to remove the 7-hydroxyl group is partially dependent on 7 alpha-hydroxysteroid dehydrogenase (HSDH) activity, which reduces 7-oxo-bile acids generated by other gut bacteria. 3 alpha-HSDH has an important enzymatic activity in the bile acid 7 alpha-dehydroxylation pathway. 12 alpha-HSDH activity has been reported for the low-activity bile acid 7 alpha-dehydroxylating bacterium Clostridium leptum; however, this activity has not been reported for high-activity bile acid 7 alpha-dehydroxylating bacteria, such as Clostridium scindens, Clostridium hylemonae, and Clostridium hiranonis. Here, we demonstrate that these strains express bile acid 12 alpha-HSDH. The recombinant enzymes were characterized from each species and shown to preferentially reduce 12-oxolithocholic acid to deoxycholic acid, with low activity against 12-oxochenodeoxycholic acid and reduced activity when bile acids were conjugated to taurine or glycine. Phylogenetic analysis suggests that 12 alpha-HSDH is widespread among Firmicutes, Actinobacteria in the Coriobacteriaceae family, and human gut Archaea. IMPORTANCE 12 alpha-HSDH activity has been established in the medically important bile acid 7 alpha-dehydroxylating bacteria C. scindens, C. hiranonis, and C. hylemonae. Experiments with recombinant 12 alpha-HSDHs from these strains are consistent with culture-based experiments that show a robust preference for 12-oxolithocholic acid over 12-oxochenodeoxycholic acid. Phylogenetic analysis identified novel members of the gut microbiome encoding 12 alpha-HSDH. Future reengineering of 12 alpha-HSDH enzymes to preferentially oxidize cholic acid may provide a means to industrially produce the therapeutic bile acid ursodeoxycholic acid. In addition, a cholic acid-specific 12 alpha-HSDH expressed in the gut may be useful for the reduction in deoxycholic acid concentration, a bile acid implicated in cancers of the gastrointestinal (GI) tract.