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Structural and biochemical characterization of 20?-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 20?-hydroxysteroid dehydrogenase (20?-HSDH) regulates cortisol side-chain cleavage by reducing the C-20 carboxyl group on cortisol, yielding 20?-dihydrocortisol. Recently, the gene encoding 20?-HSDH in Butyricicoccus desmolans ATCC 43058 was reported, and a nonredundant protein search yielded a candidate 20?-HSDH gene in Bifidobacterium adolescentis strain L2-32. B. adolescentis 20?-HSDH could regulate cortisol side-chain cleavage by limiting pro-androgen formation in bacteria such as Clostridium scindens and 21-dehydroxylation by Eggerthella lenta Here, the putative B. adolescentis 20?-HSDH was cloned, overexpressed, and purified. 20?-HSDH activity was confirmed through whole-cell and pure enzymatic assays, and it is specific for cortisol. Next, we solved the structures of recombinant 20?-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 terminus was also observed in several short-chain dehydrogenase/reductase family members. B. adolescentis strains capable of 20?-HSDH activity could alter glucocorticoid metabolism in the gut and thereby serve as potential probiotics for the management of androgen-dependent diseases.

SUBMITTER: Doden HL

PROVIDER: S-EPMC6690682 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Structural and biochemical characterization of 20β-hydroxysteroid dehydrogenase from <i>Bifidobacterium adolescentis</i> strain L2-32.

Doden Heidi L HL Pollet Rebecca M RM Mythen Sean M SM Wawrzak Zdzislaw Z Devendran Saravanan S Cann Isaac I Koropatkin Nicole M NM Ridlon Jason M JM

The Journal of biological chemistry 20190617 32

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 <i>Butyricicoccus desmo ...[more]

PMID: 31209107

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Project description:BACKGROUND: The gut microbiota plays an important role in human health and disease by acting as a metabolic organ. Metagenomic sequencing has shown how dysbiosis in the gut microbiota is associated with human metabolic diseases such as obesity and diabetes. Modeling may assist to gain insight into the metabolic implication of an altered microbiota. Fast and accurate reconstruction of metabolic models for members of the gut microbiota, as well as methods to simulate a community of microorganisms, are therefore needed. The Integrated Microbial Genomes (IMG) database contains functional annotation for nearly 4,650 bacterial genomes. This tremendous new genomic information adds new opportunities for systems biology to reconstruct accurate genome scale metabolic models (GEMs). RESULTS: Here we assembled a reaction data set containing 2,340 reactions obtained from existing genome-scale metabolic models, where each reaction is assigned with KEGG Orthology. The reaction data set was then used to reconstruct two genome scale metabolic models for gut microorganisms available in the IMG database Bifidobacterium adolescentis L2-32, which produces acetate during fermentation, and Faecalibacterium prausnitzii A2-165, which consumes acetate and produces butyrate. F. prausnitzii is less abundant in patients with Crohn's disease and has been suggested to play an anti-inflammatory role in the gut ecosystem. The B. adolescentis model, iBif452, comprises 699 reactions and 611 unique metabolites. The F. prausnitzii model, iFap484, comprises 713 reactions and 621 unique metabolites. Each model was validated with in vivo data. We used OptCom and Flux Balance Analysis to simulate how both organisms interact. CONCLUSIONS: The consortium of iBif452 and iFap484 was applied to predict F. prausnitzii's demand for acetate and production of butyrate which plays an essential role in colonic homeostasis and cancer prevention. The assembled reaction set is a useful tool to generate bacterial draft models from KEGG Orthology.

Dataset Information

Structural and biochemical characterization of 20?-hydroxysteroid dehydrogenase from Bifidobacterium adolescentis strain L2-32.

Publications

Structural and biochemical characterization of 20β-hydroxysteroid dehydrogenase from <i>Bifidobacterium adolescentis</i> strain L2-32.

Similar Datasets

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