Identification of a gene encoding a flavoprotein involved in bile acid metabolism by the human gut bacterium Clostridium scindens ATCC 35704

Spencer C. Harris, Saravanan Devendran, João M.P. Alves, Sean M. Mythen, Phillip B. Hylemon, Jason M. Ridlon

Research output: Contribution to journalArticle

Abstract

Background The multi-step bile acid 7α-dehydroxylating pathway by which a few species of Clostridium convert host primary bile acids to toxic secondary bile acids is of great importance to gut microbiome structure and host physiology and disease. While genes in the oxidative arm of the 7α-dehydroxylating pathway have been identified, genes in the reductive arm of the pathway are still obscure. Methods We identified a candidate flavoprotein-encoding gene predicted to metabolize steroids. This gene was cloned and overexpressed in E. coli and affinity purified. Reaction substrate and product were separated by thin layer chromatography and identified by liquid chromatograph mass spectrometry-ion trap-time of flight (LCMS-IT-TOF). Phylogenetic analysis of the amino acid sequence was performed. Results We report the identification of a gene encoding a flavoprotein (EDS08212.1) involved in secondary bile acid metabolism by Clostridium scindens ATCC 35704 and related species. Purified rEDS08212.1 catalyzed formation of a product from 3-dehydro-deoxycholic acid that UPLC-IT-TOF-MS analysis suggests loses 4 amu. Our phylogeny identified this gene in other bile acid 7α-dehydroxylating bacteria. Conclusions These data suggest formation of a product, 3-dehydro-4,6-deoxycholic acid, a recognized intermediate in the reductive arm of bile acid 7α-dehydroxylation pathway and the first report of a gene in the reductive arm of the bile acid 7α-dehydroxylating pathway.

Original languageEnglish (US)
Pages (from-to)276-283
Number of pages8
JournalBiochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Volume1863
Issue number3
DOIs
StatePublished - Mar 2018

Keywords

  • 7α-dehydroxylation
  • Bile acids
  • Clostridium
  • Flavoprotein

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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