Title: Structural and biochemical characterization of 20β-hydroxysteroid dehydrogenase from Bifidobacterium adolescentis strain L2-32

Abstract

Anaerobic bacteria inhabiting the human gastrointestinal tract have evolved various enzymes that modify host-derived steroids. The bacterial steroid-17,20-desmolase pathway cleaves the cortisol side chain, forming pro-androgens predicted to impact host physiology. Bacterial 20β-hydroxysteroid dehydrogenase (20β-HSDH) regulates cortisol side-chain cleavage by reducing the C-20 carboxyl group on cortisol, yielding 20β-dihydrocortisol. Recently, the gene encoding 20β-HSDH in Butyricicoccus desmolans ATCC 43058 was reported, and a nonredundant protein search yielded a candidate 20β-HSDH gene in Bifidobacterium adolescentis strain L2-32. B. adolescentis 20β-HSDH could regulate cortisol side-chain cleavage by limiting pro-androgen formation in bacteria such as Clostridium scindens and 21-dehydroxylation by Eggerthella lenta. In this work, the putative B. adolescentis 20β-HSDH was cloned, overexpressed, and purified. 20β-HSDH activity was confirmed through whole-cell and pure enzymatic assays, and it is specific for cortisol. Next, we solved the structures of recombinant 20β-HSDH in both the apo- and holo-forms at 2.0–2.2 Å resolutions, revealing close overlap except for rearrangements near the active site. Interestingly, the structures contain a large, flexible N-terminal region that was investigated by gel-filtration chromatography and CD spectroscopy. This extended N terminus is important for protein stability because deletions of varying lengths caused structural changes and reduced enzymatic activity. A nonconserved extended N terminusmore » was also observed in several short-chain dehydrogenase/reductase family members. B. adolescentis strains capable of 20β-HSDH activity could alter glucocorticoid metabolism in the gut and thereby serve as potential probiotics for the management of androgen-dependent diseases.« less

Authors:

Doden, Heidi L. ^[1]; Pollet, Rebecca M. ^[2]; Mythen, Sean M. ^[1]; Wawrzak, Zdzislaw ^[3]; Devendran, Saravanan ^[1]; Cann, Isaac ^[1]; Koropatkin, Nicole M. ^[2]; Ridlon, Jason M. ^[1]

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+ Show Author Affiliations

1. Carl R. Woese Inst. for Genomic Biology, Urbana, IL (United States); Univ. of Illinois at Urbana-Champaign, IL (United States)
2. Univ. of Michigan Medical School, Ann Arbor, MI (United States)
3. Northwestern Univ., Argonne, IL (United States). Northwestern Synchrotron Research Center–LS-CAT

Publication Date:

Mon Jun 17 00:00:00 EDT 2019

Research Org.:

Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Org.:

University of Illinois at Urbana-Champaign

OSTI Identifier:

1557315

Grant/Contract Number:  

Hatch ILLU-538-916

Resource Type:

Accepted Manuscript

Journal Name:

Journal of Biological Chemistry

Additional Journal Information:

Journal Volume: 294; Journal Issue: 32; Journal ID: ISSN 0021-9258

Publisher:

American Society for Biochemistry and Molecular Biology

Country of Publication:

United States

Language:

ENGLISH

Subject:

59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; microbiome; probiotic; androgen; glucocorticoid; oxidation–reduction (redox); 20β-hydroxysteroid dehydrogenase; Bifidobacteria; C-20 reduction; cortisol; Steroid-17,20-desmolase