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Title:Characterization of enzymatic pathways involved in cortisol and bile acid metabolism by the gut microbiota
Author(s):Mythen, Seán Martin
Advisor(s):Ridlon, Jason M
Department / Program:Animal Sciences
Discipline:Animal Sciences
Degree Granting Institution:University of Illinois at Urbana-Champaign
Bile acid
Abstract:The gut microbiota consists of a complex network of distinct bacterial taxa that together affect the physiology of the human host. Of the many facets affected by gut microbiota, their impact on the endocrine system is both significant and understudied. The diversity observed in microbial isolates and operational taxonomic units from one human gut to another is due largely in part by the strain specific proteins expressed by the microbes that inhabit them. Some of these unique proteins have evolved to biotransform host sterols which can be reabsorbed by the body and potentially affect endocrine functions. Bacterial hydroxysteroid dehydrogenases (HSDHs) have the potential to significantly alter the physicochemical properties of bile acids with implications for increased/decreased toxicity for gut bacteria and the host. We located a gene-cluster in Eggerthella CAG:298 predicted to encode three HSDHs (CDD59473, CDD59474, CDD59475), synthesized the genes for heterologous expression in E. coli and then screened bile acid substrates against the purified recombinant enzymes, revealing novel 3-, 3-, and 12-HSDHs. We also developed an enzyme-linked continuous spectrophotometric assay to quantify steroid-17,20-desmolase activity from recombinant enzymes encoded by the desA and desB genes from Clostridium scindens ATCC 35704. Steroid-17,20-desmolase is responsible for the side chain cleavage that biotransforms cortisol into 11β-hydroxyandrostenedione (11β-OHAD). The reaction performed by the steroid-17,20-desmolase appears to be regulated via pyridine nucleotide-dependent HSDHs, which are capable of converting cortisol into a form unable to be utilized by steroid-17,20-desmolase. A recently identified 20β-HSDH from Bifidobacterium adolescentis strain L2-32 has been biochemically characterized and crystallized. This 20β-HSDH has the potential to directly affect androgen formation by functioning as a metabolic rheostat controlling the steroid-17,20-desmolase activity of Clostridium scindens ATCC 35704.
Issue Date:2018-04-24
Rights Information:Copyright 2018 Seán Martin Mythen
Date Available in IDEALS:2018-09-04
Date Deposited:2018-05

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