Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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20alpha-Hydroxysteroid dehydrogenase (20 alpha-SDH) activity increases in the cycling corpus luteum of the rat, beginning at 14.00 h on the day of diestrus, but remains low in corpora lutea of pregnancy throughout the first 19 days of gestation. When cells derived from 7-day-old corpora lutea of pregnant rats were cultured for 7 or 12 days, there was a spontaneous rise in 20 alpha-SDH activity from an initial value of 0.44 +/- 0.27 to 4.1 +/- 0.7 units/mg supernatant protein. Addition of LH (NIH-S-18; 2.0 mug/ml) or prostaglandin F2alpha (2.8 X 10(-5) M) to the medium from day 4 to the end of incubation period caused a slight but significant reduction in 20 alpha-SDH activity (20%, P less than 0.05). Supplementation of the medium with ovine prolactin (HIH-P-S11; 10.0 mug/ml) from the time of seeding or from the 2nd to 4th day of culture reduced the activity of 20 alpha-SDH measured on day 12 by 61% (P less than 0.001). This finding suggests that the suppression of 20 alpha-SDH by prolactin, hitherto demonstrated only in vivo results from a direct action of the hormone on the luteal cell.
Mol Cell Endocrinol 1977 Feb
PMID:Suppression of 20 alpha-hydroxysteroid dehydrogenase activity in cultured rat luteal cells by prolactin. 83 17

20alpha-Hydroxysteroid dehydrogenase (20alpha-OH-SDH) activity was determined in the first generation corpora lutea from prepubertal rats injected with 10 I.U. of pregnant mare's serum gonadotrophin (PMSG) on day 30. The enzyme was not detectable in 1-9-day-old corpora lutea but a significant activity was seen on day 10. Enzyme activity increased during day 11 and day 12. In vivo administration of prostaglandin F2a (PGF2a) induced the enzyme in rats with corpora lutea older than 3 days. When prolactin was given concurrently with PGF2a, the corpus luteum activity of 20alpha-OH-SDH was lower than when PGF2a was given alone. It is concluded that the present "corpus luteum model" is suitable for further analysis of the cellular mechanisms of the luteolytic effect of prostaglandins (PGs) as well as of the role of gonadotrophins in the luteolytic process.
Mol Cell Endocrinol 1975 Oct
PMID:Induction by PGF2a of 20alpha-hydroxysteroid dehydrogenase in first generation corpora lutea of the rat. 119 94

Adrenocorticotropin (ACTH) is known to exert an acute effect on adrenal steroidogenesis as well as long-term effects by regulation of gene expression. In order to further study the long-term action of ACTH, guinea pig fasciculata-glomerulosa (FG) cells in primary culture were treated for up to 72 h with ACTH. The effects of this treatment on steroid secretion, enzyme activity and mRNA levels for steroid enzymes were measured. While the rate of 17-deoxy C-21 steroid secretion decreased over the 72-h period of incubation with ACTH, the 17-hydroxy C-21 steroid secretion rate remained constant for the first 24 h of incubation and declined thereafter; the rate of 4-ene C-19 steroid secretion increased over the 72-h incubation period. ACTH treatment increased 17-hydroxylase and 17,20-lyase activities and the maximal stimulation was reached after 48 h. In contrast, the activity of 21-hydroxylase (P450c21) steadily declined over the 72-h incubation period. ACTH also caused an increase in mRNA levels for P450c21, 17-hydroxylase and 17,20-lyase (P450c17), 3 beta-hydroxysteroid dehydrogenase 4-ene-5-ene-isomerase (3 beta-HSD) and cholesterol side-chain cleavage enzyme (P450scc). The maximal stimulation for the four mRNAs was observed after 18 h of incubation with ACTH, decreasing afterwards except for P450c17 mRNA levels which remained elevated over the 72-h incubation period. Despite the increase in mRNA levels for 3 beta-HSD and P450c21, no increase in their respective enzyme activities was observed and 21-hydroxylase activity even declined over the 72-h incubation period with ACTH, thus suggesting that mechanism(s) other than gene expression alone regulate steroid secretion in FG cells. In conclusion ACTH caused major changes in steroid distribution due to increased 17-hydroxylase and 17,20-lyase activities and decreased 21-hydroxylase activity in FG cells in culture. Moreover, our data revealed major differences in the induction of mRNAs for steroidogenic enzymes and their activities following ACTH treatment.
J Steroid Biochem Mol Biol 1992 Jan
PMID:Effect of ACTH on steroidogenic enzymes in guinea pig fasciculata-glomerulosa cells: changes in activity and mRNA levels. 131 Apr 15

We report here the effects of a 7-day treatment of guinea-pigs with ACTH on adrenal mRNA levels for steroid-transforming enzymes. Adrenal 3 beta-hydroxysteroid dehydrogenase 4-ene-5-ene-isomerase (3 beta-HSD), 17-hydroxylase, 17,20-lyase, 21-hydroxylase and 11-hydroxylase activities were also examined as well as plasma and adrenal steroid levels. Our data reveal that chronic ACTH-treatment stimulated all post-pregnenolone enzyme activities in glomerulosa-fasciculata cells. Plasma steroid levels increased 8 h after the last injection of ACTH and returned to the control levels 24 h later whereas, in the adrenal, the content in steroids in the group sacrificed 8 h after the last injection of ACTH were similar to the values of the control group and decreased markedly 24 h later. It is suggested that the steroid turn-over in the adrenal may be affected by the chronic ACTH-treatment. On the other hand, despite the significant stimulation in steroid-transforming enzyme activities, our data reveal that chronic ACTH administration caused a decrease in mRNA levels for P450c21 and P450c17 while P450scc, 3 beta-HSD and P450c11 remained unchanged. Taken together, these results suggest that in vivo chronic ACTH-treatment of guinea-pigs increases adrenal steroidogenic capacity by increasing steroid secretion and steroid enzyme activity. Moreover, the chronic treatment with ACTH may have a post-transcriptional effect on steroidogenic enzymes gene expression by affecting the half-life of their mRNAs.
J Steroid Biochem Mol Biol 1992 Jan
PMID:Effect of chronic ACTH treatment on guinea-pig adrenal steroidogenesis: steroid plasma levels, steroid adrenal levels, activity of steroidogenic enzymes and their steady-state mRNA levels. 131 Apr 16

Primary fetal human adrenocortical cells of definitive zone origin were transfected by electroporation with pSV3neo, a plasmid coding for SV40 T antigen and neo, which confers resistance to the antibiotic G418. The clones obtained proliferated for 30 to 40 population doublings after isolation when grown under standard medium conditions, and then entered 'crisis'. When early-passage clones were incubated with cyclic AMP (1:1 N6-monobutyryl and 8-bromo analogues), cell rounding was observed, as in primary cultures of human adrenocortical cells. As previously shown in bovine adrenocortical cells, rounding was inhibited with a monoclonal antibody against urokinase plasminogen activator but not with a monoclonal antibody against tissue plasminogen activator. The regulation of the steroidogenic pathway in clones was investigated. The effects of cyclic AMP and activation of protein kinase C were examined in cells maintained in defined medium or in the presence of serum. 17 alpha-Hydroxylase was strongly induced by cyclic AMP, as evidenced by Northern blotting and by the conversion of progesterone or 25-hydroxy-[1,2-3H]cholesterol, this induction being blocked by low concentrations of 12-O-tetradecanoylphorbol-13-acetate (TPA). Cholesterol side-chain cleavage enzyme was strongly induced by cyclic AMP, and clones also showed low activities of 21-hydroxylase and 11 beta-hydroxylase. Under all circumstances levels of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), as assessed by Northern blotting or by conversion of 25-hydroxycholesterol, were very low. 3 beta-HSD was not induced by cyclic AMP or TPA alone, but was induced by the combination of the two agents. The regulation of 17 alpha-hydroxylase and 3 beta-HSD resembles that previously described in primary cultures of human fetal adrenocortical cells. Thus, transfection with SV40 T antigen resulted in the production of clones which preserve the unique characteristics of the human adrenal cortex.
J Mol Endocrinol 1992 Aug
PMID:Expression of 17 alpha-hydroxylase and 3 beta-hydroxysteroid dehydrogenase in fetal human adrenocortical cells transfected with SV40 T antigen. 132 52

3 alpha-Hydroxysteroid dehydrogenase (3 alpha-HSD) activity has been purified to homogeneity, the enzyme is a monomer with a Mw of 32,000 Da. 3 beta-Hydroxysteroid dehydrogenase (3 beta-HSD) activity has been partially purified and has an apparent Mw of 30,000 Da. Both enzymes have the same cofactor requirements, optimal pH. However, 3 beta-HSD appeared to be an integral protein dependent on protein environment for its activity while 3 alpha-HSD activity is a protein more loosely associated to membranes.
J Steroid Biochem Mol Biol 1992 Mar
PMID:Partial purification of 3 alpha- and 3 beta-hydroxysteroid dehydrogenases from human hyperplastic prostate. Comparison between the two enzymes. 137 4

The microsomal fraction from the testes of immature pigs (less than 1 week old) contains 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta-HSD-isomerase) activities that convert dehydroepiandrosterone (DHA) to 4-androstenedione and 5,16-androstadien-3 beta-ol (andien-beta) to 4,16-androstadien-3-one (dienone). These reactions are necessary for the biosynthesis of hormonally and pheromonally active steroids. Kinetic analyses of these activities were done to determine whether they are catalysed by a single enzyme or if there is any interaction between the substrates and products of one reaction on the activity of the other enzyme. Kinetic parameters were determined and the affinities for steroid substrate were similar (7-9 mumol/l) but the Vmaxapp value for the conversion of andien-beta to dienone was 10-fold that of the DHA to 4-androstenedione reaction. In analyses of the conversion of DHA to 4-androstenedione, neither andien-beta nor dienone inhibited the reaction and especially, no effect on the Kmapp for DHA was observed which would have indicated competition between DHA and andien-beta for the same active site (Kiapp from slope and intercept replots were between 3 and 80 times the values of the kinetic constants). Similarly, DHA and 4-androstenedione had minor or negligible effects on the conversion of andien-beta to dienone (Kiapp from slope replots were the same as the Kmapp but the Kiapp from the intercept replot was 12 to 25% of the Vmaxapp). It is concluded that substrate specific 3 beta-HSD-isomerases for andien-beta and DHA exist in the immature pig testis and there is little, if any interaction between these enzymes.
J Steroid Biochem Mol Biol 1992 Sep
PMID:Kinetic evidence for separate 3 beta-hydroxysteroid dehydrogenase-isomerases in androgen and 16-androstene biosynthetic pathways in the pig testis. 138 46

3 alpha-Hydroxysteroid dehydrogenase (3 alpha-HSD) was purified greater than 500-fold from human liver cytosol. The purification was monitored using 5 beta-[3H]dihydrocortisol (5 beta-DHF) as substrate. Electrophoretically homogeneous enzyme was obtained using a procedure that involved ammonium sulfate precipitation and three successive column chromatography steps: DEAE-cellulose, hydroxylapatite and Blue-Sepharose. The enzyme is a monomer since the native molecular weight was found to be 37,000, using a calibrated Sephadex G-75 column, and the denatured subunit molecular weight was determined to be 38,500, by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The enzyme had a pI of 5.6-5.9. The 3-ketosteroids: cortisol, testosterone, progesterone and androstenedione, were not substrates for 3 alpha-HSD indicating that a saturated 4,5 double bond was required for substrate activity. The conformation at the 5 position, however, did not influence substrate activity since 5 alpha- and 5 beta-DHF and 5 alpha-dihydrotestosterone were all reduced at similar rates. The purified enzyme preferred NADPH to NADH as a cofactor and showed a broad peak of activity in the pH range of 6.8-7.4. The apparent Km for 5 beta-DHF was 18 microM. The enzyme was markedly stabilized by 50 mM phosphate buffer containing 10 to 20% glycerol at 4 degrees C. Freezing and thawing of the enzyme resulted in a large loss of activity during early stages of the purification. This is the first report of the purification to homogeneity of 3 alpha-HSD from human tissue.
J Steroid Biochem Mol Biol 1992 Oct
PMID:Purification and properties of human hepatic 3 alpha-hydroxysteroid dehydrogenase. 139 Feb 84

Primates are unique in having adrenals that secrete large amounts of the precursor sex steroids (PSS) dehydroepiandrosterone (DHEA) and especially DHEA-sulfate. The adrenal PSS require the action of 3 beta-hydroxysteroid dehydrogenase/5-ene-4 ene isomerase (3 beta-HSD), 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD), 5 alpha-reductase and/or aromatase to form the androgen dihydrotestosterone (DHT) or the estrogens 17 beta-estradiol and androst-5-ene-diol. Knowing the crucial role of 3 beta-HSD and 17 beta-HSD in sex steroid biosynthesis both in classical as well as in peripheral steroidogenic tissues, we have concentrated our efforts on the elucidation of the molecular structure of these enzyme families. We have thus characterized two types of human 3 beta-HSD cDNA clones and their corresponding genes which encode deduced proteins of 371 and 372 amino acids and share 93.5% homology. Human type I 3 beta-HSD is the almost exclusive mRNA species expressed in the placenta and skin, while human type II is the predominant mRNA species in the adrenals, ovaries and testes. We have also recently elucidated the structure of three types of rat 3 beta-HSD cDNAs which all encode a 372 amino acid protein. The predicted rat type I and II 3 beta-HSD proteins expressed adrenals, gonads and adipose tissue share 94% homology while they share 80% similarity with the liver-specific type III 3 beta-HSD. Transient expression of human type I and II as well as rat type I and II 3 beta-HSD cDNAs in HeLa human cervical carcinoma cells reveals that 3 beta-ol dehydrogenase and 5-ene-4-ene isomerase activities reside within a single protein and that these cDNAs encode functional 3 beta-HSD proteins. The expressed rat type III protein possesses a unique property catalyzing selectively the reduction of 3 beta-androstane 5 alpha-steroids such as DHT. Furthermore, we have also demonstrated by site-directed mutagenesis that the lower activity of expressed rat type II compared to rat type I 3 beta-HSD protein is due to a change of four amino acid residues potentially involved in a membrane-spanning domain. In parallel, we have characterized the complete nucleotide sequence of human 17 beta-HSD cDNA clones encoding a 327 amino acid protein as well as two in tandem 17 beta-HSD genes. Two major 17 beta-HSD mRNA species have been detected in several tissues due to a tissue-specific alternative site of initiation of transcription.(ABSTRACT TRUNCATED AT 400 WORDS)
J Steroid Biochem Mol Biol 1992 Oct
PMID:Ontogeny and subcellular localization of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) in the human and rat adrenal, ovary and testis. 139 Feb 95

The effect of long-term in vitro treatment with dexamethasone, insulin and/or LH on the 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity and the testosterone level was examined in cultures of Leydig cells from adult rats. A rapid and simple method for measuring the 3 beta-HSD activity has been developed, in which the NADH, generated by 3 beta-HSD, reduced nitroblue tetrazolium to a product with absorption maximum at 560 nm. Km for the reaction was 8.1 microM and Vmax was 12.7 nmol/min x mg protein. Addition of 0.1 or 1 microM dexamethasone for 44 h decreased the 3 beta-HSD activity to 83% and the basal testosterone level to 64% of control value after 22 and 44 h of culture. Addition of 1 nM insulin inhibited the 3 beta-HSD activity to 90% after 44 h of culture, whereas the testosterone level was increased after 3 h. Addition of 0.1 ng/ml LH did not affect the 3 beta-HSD activity in Leydig cells from adult rats. Concomitant treatment of the cells with dexamethasone and insulin inhibited the 3 beta-HSD activity to 74%, indicating an additive effect, whereas no additive effect on the testosterone level was observed. The results demonstrate that the 3 beta-HSD activity can be measured in a rapid and reliable way by measuring the reduction of nitroblue tetrazolium. Furthermore, the results suggest that dexamethasone acts on 3 beta-HSD through a mechanism different from that of insulin, as an additive effect was observed.
J Steroid Biochem Mol Biol 1992 Nov
PMID:Regulation by dexamethasone of the 3 beta-hydroxysteroid dehydrogenase activity in adult rat Leydig cells. 141 92


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