1. Structure and functional characterization of a bile acid 7α dehydratase BaiE in secondary bile acid synthesis
- Author
-
Shiva, Bhowmik, Hsien-Po, Chiu, David H, Jones, Hsiu-Ju, Chiu, Mitchell D, Miller, Qingping, Xu, Carol L, Farr, Jason M, Ridlon, James E, Wells, Marc-André, Elsliger, Ian A, Wilson, Phillip B, Hylemon, and Scott A, Lesley
- Subjects
Clostridium ,Cholic Acids ,Hydrogen Bonding ,Crystallography, X-Ray ,Hydroxylation ,Protein Structure, Secondary ,Article ,Molecular Docking Simulation ,Kinetics ,Amino Acid Substitution ,Bacterial Proteins ,Structural Homology, Protein ,Catalytic Domain ,Mutagenesis, Site-Directed ,Humans ,Hydro-Lyases ,Protein Binding - Abstract
Conversion of the primary bile acids cholic acid (CA) and chenodeoxycholic acid (CDCA) to the secondary bile acids deoxycholic acid (DCA) and lithocholic acid (LCA) is performed by a few species of intestinal bacteria in the genus Clostridium through a multistep biochemical pathway that removes a 7α-hydroxyl group. The rate-determining enzyme in this pathway is bile acid 7α-dehydratase (baiE). In this study, we report crystal structures of apo-BaiE and its putative product-bound (3-oxo-Δ4,6- lithocholyl-Coenzyme A (CoA)) complex. BaiE is a trimer with a twisted α+β barrel fold with similarity to the Nuclear Transport Factor 2 (NTF2) superfamily. Tyr30, Asp35 and His83 form a catalytic triad that is conserved across this family. Site-directed mutagenesis of BaiE from Clostridium scindens VPI 12708 confirmed that these residues are essential for catalysis and also confirmed the importance of other conserved residues, Tyr54 and Arg146, which are involved in substrate binding and affect catalytic turnover. Steady state kinetic studies revealed that the BaiE homologs are able to turn over 3-oxo-Δ4-bile acid and CoA-conjugated 3-oxo-Δ4-bile acid substrates with comparable efficiency questioning the role of CoA-conjugation in the bile acid metabolism pathway.
- Published
- 2015