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Expression in Escherichia coli and characterization of a bile acid-inducible 3α-hydroxysteroid dehydrogenase from Eubacterium sp. strain VPI 12708

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Abstract

We have previously cloned and sequenced three members of a bile acid-inducible gene family from Eubacterium sp. strain VPI 12708 that encode 27,000-M r polypeptides. Two copies of these genes (baiA1 and baiA3) are identical, while the third copy (baiA2) encodes a polypeptide sharing 92% amino acid identity with the baiA1 and baiA3 gene products. We have overexpressed the baiA1 gene in Escherichia coli and analyzed the expressed activity. Thin-layer chromatography of 14C-labeled bile acid products from reactions using cell-free extracts revealed a 3α-hydroxysteroid dehydrogenase activity for the BaiA1 protein. The BaiA1 protein could utilize both NAD+ and NADP+, and the preferred steroid substrate was the cholyl-coenzyme A conjugate rather than free cholic acid. These results show that the BaiA proteins are novel 3α-hydroxysteroid dehydrogenases.

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Authors and Affiliations

  1. Department of Microbiology and Immunology, Medical College of Virginia, Virginia Commonwealth University, 23298-0678, Richmond, VA, USA

    Darrell H. Mallonee, Mark A. Lijewski & Phillip B. Hylemon

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  1. Darrell H. Mallonee
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  2. Mark A. Lijewski
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  3. Phillip B. Hylemon
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Mallonee, D.H., Lijewski, M.A. & Hylemon, P.B. Expression in Escherichia coli and characterization of a bile acid-inducible 3α-hydroxysteroid dehydrogenase from Eubacterium sp. strain VPI 12708. Current Microbiology 30, 259–263 (1995). https://doi.org/10.1007/BF00295498

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