Clostridium scindens baiCD and baiH genes encode stereo-specific 7alpha/7beta-hydroxy-3-oxo-delta4-cholenoic acid oxidoreductases.

Academic Article

Abstract

  • Secondary bile acids, formed by intestinal bacteria, are suggested to play a significant role in cancers of the gastrointestinal tract in humans. Bile acid 7alpha/beta-dehydroxylation is carried out by a few species of intestinal clostridia which harbor a multi-gene bile acid inducible (bai) operon. Several genes encoding enzymes in this pathway have been cloned and characterized. However, no gene product(s) has yet been assigned to the production of 3-oxo-Delta4-cholenoic acid intermediates of cholic acid (CA), chenodeoxycholic acid (CDCA) or ursodeoxycholic acid (UDCA). We previously reported that the baiH gene encodes an NADH:flavin oxidoreductase (NADH:FOR); however, the role of this protein in bile acid 7-dehydroxylation is unclear. Homology searches and secondary structural alignments suggest this protein to be similar to flavoproteins which reduce alpha/beta-unsaturated carbonyl compounds. The baiH gene product was expressed in Escherichia coli, purified and discovered to be a stereo-specific NAD(H)-dependent 7beta-hydroxy-3-oxo-Delta4-cholenoic acid oxidoreductase. Additionally, high sequence similarity between the baiH and baiCD gene products suggests the baiCD gene may encode a 3-oxo-Delta4-cholenoic acid oxidoreductase specific for CDCA and CA. We tested this hypothesis using cell extracts prepared from E. coli overexpressing the baiCD gene and discovered that it encodes a stereo-specific NAD(H)-dependent 7alpha-hydroxy-3-oxo-Delta4-cholenoic acid oxidoreductase.
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    Keywords

  • Amino Acid Sequence, Carboxylic Acids, Catalysis, Chromatography, Thin Layer, Clostridium, Conserved Sequence, Molecular Sequence Data, Oxidoreductases, Protein Structure, Secondary, Recombinant Proteins, Sequence Alignment, Stereoisomerism, Substrate Specificity
  • Digital Object Identifier (doi)

    Author List

  • Kang D-J; Ridlon JM; Moore DR; Barnes S; Hylemon PB
  • Start Page

  • 16
  • End Page

  • 25
  • Volume

  • 1781
  • Issue

  • 1-2