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Jason M. Ridlon - One of the best experts on this subject based on the ideXlab platform.
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Gut feelings about bacterial steroid-17,20-Desmolase.
Molecular and cellular endocrinology, 2021Co-Authors: Heidi L. Doden, Jason M. RidlonAbstract:Abstract Advances in technology are only beginning to reveal the complex interactions between hosts and their resident microbiota that have co-evolved over centuries. In this review, we present compelling evidence that implicates the host-associated microbiome in the generation of 11β-hydroxyandrostenedione, leading to the formation of potent 11-oxy-androgens. Microbial steroid-17,20-Desmolase cleaves the side-chain of glucocorticoids (GC), including cortisol (and its derivatives of cortisone, 5α-dihydrocortisol, and also (allo)- 3α, 5α-tetrahydrocortisol), but not 3α-5β-tetrahydrocortisol) and drugs (prednisone and dexamethasone). In addition to side-chain cleavage, we discuss the gut microbiome’s robust potential to transform a myriad of steroids, mirroring much of the host’s metabolism. We also explore the overlooked role of intestinal steroidogenesis and efflux pumps as a potential route for GC transport into the gut. Lastly, we propose several health implications from microbial steroid-17,20-Desmolase function, including aberrant mineralocorticoid, GC, and androgen receptor signaling in colonocytes, immune cells, and prostate cells, which may exacerbate disease states.
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The desA and desB genes from Clostridium scindens ATCC 35704 encode steroid-17,20-Desmolase
Journal of lipid research, 2018Co-Authors: Saravanan Devendran, Sean M. Mythen, Jason M. RidlonAbstract:Clostridium scindens is a gut microbe capable of removing the side-chain of cortisol, forming 11β-hydro-xyandrostenedione. A cortisol-inducible operon (desABCD) was previously identified in C. scindens ATCC 35704 by RNA-Seq. The desC gene was shown to encode a cortisol 20α-hydroxysteroid dehydrogenase (20α-HSDH). The desD encodes a protein annotated as a member of the major facilitator family, predicted to function as a cortisol transporter. The desA and desB genes are annotated as N-terminal and C-terminal transketolases, respectively. We hypothesized that the DesAB forms a complex and has steroid-17,20-Desmolase activity. We cloned the desA and desB genes from C. scindens ATCC 35704 in pETDuet for overexpression in Escherichia coli The purified recombinant DesAB was determined to be a 142 ± 5.4 kDa heterotetramer. We developed an enzyme-linked continuous spectrophotometric assay to quantify steroid-17,20-Desmolase. This was achieved by coupling DesAB-dependent formation of 11β-hydroxyandrostenedione with the NADPH-dependent reduction of the steroid 17-keto group by a recombinant 17β-HSDH from the filamentous fungus, Cochliobolus lunatus The pH optimum for the coupled assay was 7.0 and kinetic constants using cortisol as substrate were Km of 4.96 ± 0.57 µM and kcat of 0.87 ± 0.076 min-1 Substrate-specificity studies revealed that rDesAB recognized substrates regardless of 11β-hydroxylation, but had an absolute requirement for 17,21-dihydroxy 20-ketosteroids.
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Identification and characterization of a 20β-HSDH from the anaerobic gut bacterium Butyricicoccus desmolans ATCC 43058
Journal of lipid research, 2017Co-Authors: Saravanan Devendran, Celia Méndez-garcía, Jason M. RidlonAbstract:Members of the gastrointestinal microbiota are known to convert glucocorticoids to androstanes, which are subsequently converted to potent androgens by other members of the gut microbiota or host tissues. Butyricicoccus desmolans and Clostridium cadaveris have previously been reported for steroid-17,20-Desmolase and 20β-hydroxysteroid dehydrogenase (HSDH) activities that are responsible for androstane formation from cortisol; however, the genes encoding these enzymes have yet to be reported. In this work, we identified and located a gene encoding 20β-HSDH in both B. desmolans and C. cadaveris The 20β-HSDH of B. desmolans was heterologously overexpressed and purified from Escherichia coli The enzyme was determined to be a homotetramer with subunit molecular mass of 33.8 ± 3.7 kDa. The r20β-HSDH displayed pH optimum in the reductive direction at pH 9.0 and in the oxidative direction at pH 7.0-7.5 with (20β-dihydro)cortisol and NAD(H) as substrates. Cortisol is the preferred substrate with Km , 0.80 ± 0.06 μM; Vmax , 30.36 ± 1.97 μmol·min-1; Kcat , 607 ± 39 μmol·μM-1·min-1; Kcat /Km , 760 ± 7.67. Phylogenetic analysis of the 20β-HSDH from B. desmolans suggested that the 20β-HSDH is found in several Bifidobacterium spp, one of which was shown to express 20β-HSDH activity. Notably, we also identified a novel steroid-17,20-Desmolase-elaborating bacterium, Propionimicrobium lymphophilum, a normal inhabitant of the urinary tract.
M Zachmann - One of the best experts on this subject based on the ideXlab platform.
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Defects in steroidogenic enzymes. Discrepancies between clinical steroid research and molecular biology results.
The Journal of steroid biochemistry and molecular biology, 1995Co-Authors: M ZachmannAbstract:Molecular biology has clarified the understanding of steroidogenic enzyme genetics. Nevertheless, there are discrepancies between fundamental and clinical experience. (1) Why do patients with "pure" 17 alpha-hydroxylase or 17,20-Desmolase deficiency exist, when one cytochrome regulates both steps? A case of interest is discussed, who had "pure" 17,20-Desmolase deficiency until adolescence, but additional 17 alpha-hydroxylase deficiency thereafter. (2) In 11 beta-hydroxylase deficiency, it was puzzling to find 18-hydroxylated compounds, and, in isolated hypoaldosteronism, normal cortisol, since 11 beta- and 18-hydroxylation were thought to be regulated together. This has now been explained by differences in the fasciculata and glomerulosa. The occurrence of 11 beta-hydroxylase deficiency of 17-hydroxylated steroids only, however, remains enigmatic. (3) 3 beta-Hydroxysteroid dehydrogenase deficiency does not only seem to exist in classic (mutations of type II gene), but also in late-onset cases. In them, no molecular basis could be found. (4) Also, in cholesterol side-chain cleavage, there is an inequity: while evidently one cytochrome regulates 20- and 22-hydroxylation, pregnenolone is formed when 20 alpha OH-cholesterol, but not when cholesterol, is added to adrenal tissue of deficient patients. Other factors (promoters, fusion proteins, adrenodoxin, cAMP-dependent expression of genes, and/or proteases), or hormonal replacement in patients may be responsible for these discrepancies.
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Endocrine findings in male pseudohermaphroditism.
European journal of pediatrics, 1993Co-Authors: M ZachmannAbstract:Recent discoveries in molecular biology have much clarified the regulation and function of steroid converting enzymes. Most progress has been made in the area of cytochromes, which regulate the side chain cleavage of cholesterol (P-450 SCC) and the 17 alpha-hydroxylase- and 17,20-Desmolase (or 17,20-lyase) activities (P-450 17 alpha), as well as in 3 beta-hydroxysteroid dehydrogenase. Nevertheless, there are some discrepancies between fundamental knowledge and clinical experience, which are difficult to understand: why is it possible, e.g., that cases with "pure" 17 alpha-hydroxylase or 17,20-Desmolase deficiency exist, when there is only one cytochrome regulating both steps? After a brief review of clinical and biochemical findings in the various defects of testosterone biosynthesis, a case is discussed which is of interest in this respect.
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conversion from pure 17 20 Desmolase to combined 17 20 Desmolase 17α hydroxylase deficiency with age
European Journal of Endocrinology, 1992Co-Authors: M Zachmann, B. Kempken, B. Manella, Elizabeth NavarroAbstract:A previously described phenotypically female 46,XY-individual with 17,20-Desmolase deficiency was followed from adolescence to adulthood. While at age 16.9 years her 17 alpha-hydroxylating capacity was normal and steroids not hydroxylated in position 17 low, decreasing 17 alpha-hydroxylated and increasing unhydroxylated steroids were observed thereafter up to the age of 25.2 years. Simultaneously with the steroid changes, previously normal blood pressure (110/80 mmHg) increased (180/130 mmHg). Since 17 alpha-hydroxylation and 17,20-Desmolase activity are regulated by one cytochrome P450-17 alpha, it is concluded that, in this same patient, 17 alpha-hydroxylase activity was normal during childhood and adolescence, but decreased in the young adult. 17,20-Desmolase activity, by contrast, was completely absent already in the fetus, causing absence of masculinization. The factors modulating this difference in cytochrome P450-17 alpha activity with age are as yet unknown.
Milo Zachmann - One of the best experts on this subject based on the ideXlab platform.
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Endocrine findings in male pseudohermaphroditism
European Journal of Pediatrics, 1993Co-Authors: Milo ZachmannAbstract:Recent discoveries in molecular biology have much clarified the regulation and function of steroid converting enzymes. Most progress has been made in the area of cytochromes, which regulate the side chain cleavage of cholesterol (P-450 SCC) and the 17α-hydroxylase- and 17, 20-Desmolase (or 17, 20-lyase) activities (P-450 17α), as well as in 3β-hydroxysteroid dehydrogenase. Nevertheless, there are some discrepancies between fundamental knowledge and clinical experience, which are difficult to understand: why is it possible, e.g., that cases with “pure” 17α-hydroxylase or 17,20-Desmolase deficiency exist, when there is only one cytochrome regulating both steps? After a brief review of clinical and biochemical findings in the various defects of testosterone biosynthesis, a case is discussed which is of interest in this respect.
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Conversion from pure 17,20-Desmolase- to combined 17,20-Desmolase/17α-hydroxylase deficiency with age
Acta endocrinologica, 1992Co-Authors: Milo Zachmann, B. Kempken, B. Manella, Elizabeth NavarroAbstract:A previously described phenotypically female 46,XY-individual with 17,20-Desmolase deficiency was followed from adolescence to adulthood. While at age 16.9 years her 17 alpha-hydroxylating capacity was normal and steroids not hydroxylated in position 17 low, decreasing 17 alpha-hydroxylated and increasing unhydroxylated steroids were observed thereafter up to the age of 25.2 years. Simultaneously with the steroid changes, previously normal blood pressure (110/80 mmHg) increased (180/130 mmHg). Since 17 alpha-hydroxylation and 17,20-Desmolase activity are regulated by one cytochrome P450-17 alpha, it is concluded that, in this same patient, 17 alpha-hydroxylase activity was normal during childhood and adolescence, but decreased in the young adult. 17,20-Desmolase activity, by contrast, was completely absent already in the fetus, causing absence of masculinization. The factors modulating this difference in cytochrome P450-17 alpha activity with age are as yet unknown.
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Recent aspects of steroid biosynthesis in male sex differentiation. Clinical studies.
Hormone research, 1992Co-Authors: Milo ZachmannAbstract:Recent discoveries in molecular biology have much clarified the regulation and function of steroid-converting enzymes. Most progress has been made in the area of cytochromes, which regulate the side chain cleavage of cholesterol (P-450 SCC) and the 17α-hydroxylase and 17,20-Desmolase (or 17,20-lyase) activities (P-450 17α), as well as in 3β-hydroxysteroid dehydrogenase. Nevertheless, there are some discrepancies between fundamental knowledge and clinical experience, which are difficult to understand: why is it for example possible that cases with ‘pure’ 17α-hydroxylase or 17,20-Desmolase deficiency exist, when there is only one cytochrome regulating both steps? After a brief review of clinical and biochemical findings in the various defects of testosterone biosynthesis, a case is discussed, which is of interest in this respect. This XY patient with female external genitalia, who has been shown to have compound heterozygous mutations, had ‘pure’ 17,20-Desmolase deficiency up to adolescence, but additional 17α-hydroxylase deficiency with hypertension developed thereafter. From this observation, it has to be concluded that as yet unknown, possibly age-dependent modulating factors exist, which influence the activity of the cytochrome. Also the estrogen replacement given to the patient might have played a role in this change.
Dieter Engelhardt - One of the best experts on this subject based on the ideXlab platform.
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Different inhibitory effect of etomidate and ketoconazole on the human adrenal steroid biosynthesis.
The Clinical Investigator, 1993Co-Authors: Matthias M. Weber, J. Lang, F. Abedinpour, K. Zeilberger, B. C. Adelmann, Dieter EngelhardtAbstract:The narcotic agent etomidate and the antimycotic drug ketoconazole are known to block steroid biosynthesis in man. To study the different effects of these imidazole derivatives on human adrenal steroid biosynthesis we incubated slices of human adrenal glands with 3H-labeled precursors and increasing concentrations of etomidate or ketoconazole (0-2000 μM). After extraction the labeled metabolites were separated by thin-layer chromatography and quantified by scintillation counting. Etomidate inhibited most potently 11β-hydroxylase activity by suppressing the formation of corticosterone from 11-deoxycorticosterone to 1 % of control [50% inhibitory concentration (IC50) 0.03 μM] while ketoconazole suppressed 11β-hy-droxylase to only 39% of control activity (IC50 15 μM). Ketoconazole however, most potently blocked the conversion of 17α-hydroxy-proges-terone to androstenedione by C17,20-Desmolase to about 15% of control activity (IC50 1 μM) while etomidate showed a much weaker effect on this enzyme with a suppression to 50% of C17,20-Desmolase control activity at a concentration of 380 μM. Both imidazole drugs showed a similar strong inhibitory effect on the activity of 17α-hy-droxylase (IC50 6-18 μM) and 16α-hydroxylase (IC50 4–8 μM) and did not affect 21-hydroxylase. These in vitro data indicate a predominant inhibitory effect of etomidate on corticosteroid biosynthesis by relative selective inhibition of 11β-hydroxylase and of ketoconazole on the adrenal androgen biosynthesis by a predominant inhibition of C17,20-Desmolase. This differential inhibitory effect of etomidate and ketoconazole on human steroid biosynthesis may be of clinical importance for a possible therapeutic use of these imidazole derivatives in endocrine disorders.
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Effect of ketoconazole on human ovarian C17,20-Desmolase and aromatase.
The Journal of steroid biochemistry and molecular biology, 1991Co-Authors: Matthias M. Weber, B. C. Adelmann, A. Will, Dieter EngelhardtAbstract:Ketoconazole, an imidazole antimycotic drug, inhibits steroid biosynthesis in adrenal and testicular tissue by blocking cytochrome P-450 dependent enzymes. To study the effect of ketoconazole on steroid biosynthesis in the human ovary we incubated human ovarian tissue (mainly theca cells) or granulosa cells with radiolabeled precursors and increasing concentrations of ketoconazole. After incubation, steroids were extracted and separated by thin layer chromatography (TLC). Activity of C17,20-Desmolase and aromatase was estimated by measuring the amount of their radioactive products with liquid scintillation counting. After incubation of ovarian tissue with [3H]17-hydroxyprogesterone the production of [3H]androstenedione was reduced by increasing concentrations of ketoconazole (0–200 μM) to a minimum of 31% of basal production. This indicates a strong inhibition of ovarian C17,20-Desmolase by ketoconazole with a 50% inhibiting concentration (IC50) of 23 μM. After incubation of human granulosa cells with ketoconazole (0–2000 μM) and [3H]androstenedione the production of [3H]estrone and [3H]estradiol was suppressed to minimally 37 and 35% of basal values, indicating a significant inhibition of ovarian aromatase. IC50-values were 105 μM ketoconazole for estradiol and 130 μM for estrone. In conclusion, ketoconazole was shown to inhibit human ovarian C17,20-Desmolase and aromatase in vitro. As in human adrenals and testes ovarian C17,20-Desmolase seems to be most sensitive to the inhibitory effect of ketoconazole.
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The influence of ketoconazole on human adrenal steroidogenesis: incubation studies with tissue slices.
Clinical endocrinology, 1991Co-Authors: Dieter Engelhardt, Matthias M. Weber, F. Abedinpour, T. Miksch, C. JaspersAbstract:OBJECTIVE The influence of ketoconazole on the various enzymes of human adrenal steroid biosynthesis was examined in vitro. MEASUREMENTS After incubation of human adrenal tissue slices with labelled precursors and ketoconazole (0-2000 microM), radioactive metabolites were separated by thin-layer chromatography and quantified by liquid scintillation counting. Enzyme activity was assessed by measuring conversion of tritium-labelled precursors to products. RESULTS In vitro, ketoconazole showed a significant inhibition on the following adrenal enzyme systems (with decreasing activity): C17,20-Desmolase (IC50 2 microM), 16 alpha-hydroxylase (IC50 9 microM), 17 alpha-hydroxylase (IC50 18 microM), 18-hydroxylase (IC50 28 microM), and 11 beta-hydroxylase (IC50 35 microM). In the tested concentrations ketoconazole had no inhibitory effect on the 21-hydroxylase, the 3 beta-hydroxysteroid dehydrogenase and the 20-hydroxysteroid dehydrogenase component of the C17,20-Desmolase enzyme system. CONCLUSIONS The data are in accordance with clinical findings where a strong suppression of serum androgen levels by relatively selective inhibition of C17, 20-Desmolase has been assumed. The predominant blocking effect of ketoconazole on adrenal as well as on gonadal androgen biosynthesis might be of clinical benefit in the management of hyperandrogenic states.
Saravanan Devendran - One of the best experts on this subject based on the ideXlab platform.
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Bacterial steroid-17,20-Desmolase is a taxonomically rare enzymatic pathway that converts prednisone to 1,4-androstanediene-3,11,17-trione, a metabolite that causes proliferation of prostate cancer cells.
The Journal of steroid biochemistry and molecular biology, 2019Co-Authors: Joe L. Rowles, Saravanan Devendran, Hans Müller Paul, João M. P. Alves, Camdon Yemm, Patricia M. Wolf, Matthew E. Hudson, David J. Morris, John W. ErdmanAbstract:Abstract The adrenal gland has traditionally been viewed as a source of “weak androgens”; however, emerging evidence indicates 11-oxy-androgens of adrenal origin are metabolized in peripheral tissues to potent androgens. Also emerging is the role of gut bacteria in the conversion of C21 glucocorticoids to 11-oxygenated C19 androgens. Clostridium scindens ATCC 35,704 is a gut microbe capable of converting cortisol into 11-oxy-androgens by cleaving the side-chain. The desA and desB genes encode steroid-17,20-Desmolase. Our prior study indicated that the urinary tract bacterium, Propionimicrobium lymphophilum ACS-093-V-SCH5 encodes desAB and converts cortisol to 11β-hydroxyandrostenedione. We wanted to determine how widespread this function occurs in the human microbiome. Phylogenetic and sequence similarity network analyses indicated that the steroid-17,20-Desmolase pathway is taxonomically rare and located in gut and urogenital microbiomes. Two microbes from each of these niches, C. scindens and Propionimicrobium lymphophilum, respectively, were screened for activity against endogenous (cortisol, cortisone, and allotetrahydrocortisol) and exogenous (prednisone, prednisolone, dexamethasone, and 9-fluorocortisol) glucocorticoids. LC/MS analysis showed that both microbes were able to side-chain cleave all glucocorticoids, forming 11-oxy-androgens. Pure recombinant DesAB from C. scindens showed the highest activity against prednisone, a commonly prescribed glucocorticoid. In addition, 0.1 nM 1,4-androstadiene-3,11,17-trione, bacterial side-chain cleavage product of prednisone, showed significant proliferation relative to vehicle in androgen-dependent growth LNCaP prostate cancer cells after 24 h (2.3 fold; P
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The desA and desB genes from Clostridium scindens ATCC 35704 encode steroid-17,20-Desmolase
Journal of lipid research, 2018Co-Authors: Saravanan Devendran, Sean M. Mythen, Jason M. RidlonAbstract:Clostridium scindens is a gut microbe capable of removing the side-chain of cortisol, forming 11β-hydro-xyandrostenedione. A cortisol-inducible operon (desABCD) was previously identified in C. scindens ATCC 35704 by RNA-Seq. The desC gene was shown to encode a cortisol 20α-hydroxysteroid dehydrogenase (20α-HSDH). The desD encodes a protein annotated as a member of the major facilitator family, predicted to function as a cortisol transporter. The desA and desB genes are annotated as N-terminal and C-terminal transketolases, respectively. We hypothesized that the DesAB forms a complex and has steroid-17,20-Desmolase activity. We cloned the desA and desB genes from C. scindens ATCC 35704 in pETDuet for overexpression in Escherichia coli The purified recombinant DesAB was determined to be a 142 ± 5.4 kDa heterotetramer. We developed an enzyme-linked continuous spectrophotometric assay to quantify steroid-17,20-Desmolase. This was achieved by coupling DesAB-dependent formation of 11β-hydroxyandrostenedione with the NADPH-dependent reduction of the steroid 17-keto group by a recombinant 17β-HSDH from the filamentous fungus, Cochliobolus lunatus The pH optimum for the coupled assay was 7.0 and kinetic constants using cortisol as substrate were Km of 4.96 ± 0.57 µM and kcat of 0.87 ± 0.076 min-1 Substrate-specificity studies revealed that rDesAB recognized substrates regardless of 11β-hydroxylation, but had an absolute requirement for 17,21-dihydroxy 20-ketosteroids.
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Identification and characterization of a 20β-HSDH from the anaerobic gut bacterium Butyricicoccus desmolans ATCC 43058
Journal of lipid research, 2017Co-Authors: Saravanan Devendran, Celia Méndez-garcía, Jason M. RidlonAbstract:Members of the gastrointestinal microbiota are known to convert glucocorticoids to androstanes, which are subsequently converted to potent androgens by other members of the gut microbiota or host tissues. Butyricicoccus desmolans and Clostridium cadaveris have previously been reported for steroid-17,20-Desmolase and 20β-hydroxysteroid dehydrogenase (HSDH) activities that are responsible for androstane formation from cortisol; however, the genes encoding these enzymes have yet to be reported. In this work, we identified and located a gene encoding 20β-HSDH in both B. desmolans and C. cadaveris The 20β-HSDH of B. desmolans was heterologously overexpressed and purified from Escherichia coli The enzyme was determined to be a homotetramer with subunit molecular mass of 33.8 ± 3.7 kDa. The r20β-HSDH displayed pH optimum in the reductive direction at pH 9.0 and in the oxidative direction at pH 7.0-7.5 with (20β-dihydro)cortisol and NAD(H) as substrates. Cortisol is the preferred substrate with Km , 0.80 ± 0.06 μM; Vmax , 30.36 ± 1.97 μmol·min-1; Kcat , 607 ± 39 μmol·μM-1·min-1; Kcat /Km , 760 ± 7.67. Phylogenetic analysis of the 20β-HSDH from B. desmolans suggested that the 20β-HSDH is found in several Bifidobacterium spp, one of which was shown to express 20β-HSDH activity. Notably, we also identified a novel steroid-17,20-Desmolase-elaborating bacterium, Propionimicrobium lymphophilum, a normal inhabitant of the urinary tract.