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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD) is a microsomal enzyme complex responsible for the interconversion of active 11-hydroxy glucocorticoids to inactive 11-oxo metabolites. It has long been controversially discussed whether 11-dehydrogenation and 11-oxoreduction are catalysed by a single bidirectional enzyme or if the 11 beta-HSD system comprises 2 kinetically distinct microsomal enzyme activities, 11-dehydrogenase and 11-oxoreductase. However, 11-oxoreduction of homogeneously purified 11 beta-HSD could not be demonstrated under in vitro conditions until today. We have purified 11 beta-HSD from mouse liver microsomes to homogeneity by a purification method which affords a gentle membrane protein solubilization as well as providing a favourable detergent surrounding during the various chromatographic steps. Following 11-dehydrogenation of corticosterone and 11-oxoreduction of dehydrocorticosterone simultaneously throughout the entire purification procedure we could demonstrate that 11 beta-HSD retains both oxidative and reductive activities in almost the same ratio, which is also true for the homogeneously purified enzyme. Deducing from the coincidentally increasing specific activities of 11-dehydrogenation and 11-oxoreduction the conclusion can be drawn that both activities reside within the same protein. Furthermore, in addition to NADP(H) also NAD(H) can serve as cosubstrate, which is mainly true for the oxidative direction. In conclusion, our results provide evidence that the oxidative and reductive behaviour of 11 beta-HSD can be explained by the concept of a unique, reversible oxidoreductase thus disproving the two enzyme theory.
J Steroid Biochem Mol Biol 1994 Feb
PMID:The purification of 11 beta-hydroxysteroid dehydrogenase from mouse liver microsomes. 751 8

Pathways of testosterone metabolism in tissue slices and cell suspensions of human benign hyperplastic prostate (BPH) tissue and human prostate cancer cell lines (DU145, HPC-36M, PC-3/MA2 and LNCaP) were investigated. Thin layer chromatography analysis was used to identify the following tritiated metabolites: testosterone, 5 alpha-dihydrostestosterone (DHT), 5 alpha-androstane-3 alpha/3 beta-17 beta-diol (androstanediols), 4-androstene-3,17-dione (androstenedione) and 5 alpha-androstanedione. The predominant pathway for testosterone metabolism in BPH was via 5 alpha-reductase producing 5 alpha-dihydrotestosterone (71% and 75% total metabolites in slices and suspensions incubated for 24 h, respectively). The cancer cell lines DU145 and HPC-36M resembled BPH by metabolizing testosterone predominantly to DHT (68% and 82% total metabolites, respectively), although the rate of metabolism was much lower in the cell lines (0.099 and 0.05 pmol testosterone/mg protein/h in DU145 and HPC-36M) compared to the BPH cell suspensions (6.4 pmol testosterone/mg protein/h). In contrast, PC-3/MA2 contained high 17 beta-HSD activity forming large amounts of 4-androstene-3,17-dione (84% total metabolites), converting testosterone at a rate faster (12.8 pmol testosterone/mg protein/h) than the BPH cell suspensions. LNCaP rapidly converted testosterone exclusively to a glucuronide conjugate (7.4 pmol testosterone/mg protein/h), although after incubation with [3H]-4-androstene-3,17-dione, 5 alpha-reductase activity was demonstrated. LNCaP was the only cell line whose growth and colony-forming ability was stimulated by testosterone and DHT. BPH and all the cell lines tested had 5 alpha-reductase activity, but only the prostate tissue and the cell lines DU145 and HPC-36M converted testosterone predominantly to DHT.
J Steroid Biochem Mol Biol 1994 Aug
PMID:Comparison of testosterone metabolism in benign prostatic hyperplasia and human prostate cancer cell lines in vitro. 751 39

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD), responsible for the interconversion of hormonally active cortisol to inactive cortisone, dictates specificity for the mineralocorticoid receptor (MR) in the distal nephron and colon. Two isoforms of human 11 beta-HSD have been cloned, an NADP(H)-dependent (type 1) dehydrogenase/oxo-reductase enzyme, and a high-affinity NAD-dependent (type 2) unidirectional dehydrogenase. Using the reverse-transcriptase polymerase chain reaction (RT-PCR) amplification of RNA extracted from human adult tissues, type 1 11 beta-HSD mRNA was found in decidua, placenta, liver, lung, spleen, kidney medulla, cerebellum and pituitary, but was absent in kidney cortex, sigmoid and rectal colon, salivary gland and thyroid. In contrast, type 2 11 beta-HSD mRNA was found only in placenta and in the classical mineralocorticoid target tissues, kidney cortex, kidney medulla, sigmoid and rectal colon, salivary gland, and colonic epithelial cell lines (AAC1 and RGC28). In situ hybridization studies of renal cortex, cortico-medullary junction and medulla using a 35S-labeled antisense cRNA probe for type 2 human 11 beta-HSD, revealed specific localization of type 2 11 beta-HSD mRNA expression exclusively to renal cortical and medullary collecting ducts. Type 1 and type 2 isoforms of human 11 beta-HSD are expressed in a distinct tissue-specific fashion, in keeping with the proposed differences in their physiological roles. Type 2 11 beta-HSD is found predominantly in mineralocorticoid target tissues where it serves to protect the MR in an autocrine fashion.
Mol Cell Endocrinol 1995 Apr 28
PMID:Detection of human 11 beta-hydroxysteroid dehydrogenase isoforms using reverse-transcriptase-polymerase chain reaction and localization of the type 2 isoform to renal collecting ducts. 754 19

The cellular localization of 11 beta-hydroxysteroid dehydrogenase 2 (11 beta-HSD2) gene expression in the ovine adrenal gland was determined by in situ hybridization histochemistry. 11 beta-HSD2 mRNA was localized exclusively to the adrenal cortex of the adult sheep, and within the cortex the mRNA was highly expressed in the zona fasciculata and zona reticularis with relatively low expression in the zona glomerulosa. Radiometric conversion assay using adrenal cortical tissues revealed extremely high levels of 11 beta-HSD activity which was characteristic of 11 beta-HSD2 in that it was NAD-dependent and displayed a Km for cortisol of 41 +/- 4 nM. This indicates that 11 beta-HSD2 mRNA within the ovine adrenal gland is translated and functional with respect to enzymatic activity. In marked contrast, 11 beta-HSD1 mRNA was undetectable in either the cortex or medulla of adult sheep adrenal glands. In conclusion, we have demonstrated, for the first time, the zonal localization of 11 beta-HSD2 mRNA and the presence of 11 beta-HSD2 activity in the adult sheep adrenal cortex. The adrenal 11 beta-HSD2 may function to (1) regulate the rate of cortisol secretion by adrenocortical cells; (2) protect these cells from high levels of locally produced glucocorticoids; and/or (3) provide an important source of circulating cortisone, which can be activated by the action of 11 beta-HSD1 reductase in organs such as the liver.
Mol Cell Endocrinol 1995 Jun
PMID:Cellular localization of 11 beta-hydroxysteroid dehydrogenase 2 gene expression in the ovine adrenal gland. 755 71

The cDNA coding for pig testicular 3 alpha/beta (20 beta)-hydroxysteroid dehydrogenase was expressed in Escherichia coli by placing it under the control of an isopropylthiogalactoside (IPTG) inducible tac promoter. Production of 3 alpha/beta (20 beta)-HSD was demonstrated by Western blotting and by catalytic activity with 5 alpha-dihydrotestosterone as a substrate for 3 alpha/beta-HSD, and progesterone and 17 alpha-hydroxyprogesterone as substrates for 20 beta-HSD in the presence of NADPH. The 3 alpha/beta (20 beta)-HSD enzyme was detected in a soluble fraction of the lysate of E. coli added to IPTG to induce the synthesis of the protein. Its molecular weight was estimated to be 30.5 kDa by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Recombinant 3 alpha/beta (20 beta)-HSD was purified to apparent homogeneity as determined by SDS-PAGE by column chromatography using DEAE-cellulose. The purified enzyme reduced not only steroids but also prostaglandins and other carbonyl compounds including aldehydes, ketones and quinones as demonstrated in native enzymes purified from pig testes. The amino terminus of the purified enzyme was serine which was coded next to the ATG start codon, and the sequence of the amino terminal 24 residues was identical with the coding amino acid in the cDNA; whereas, the amino terminus of the native 3 alpha/beta (20 beta)-HSD was not detected suggesting that the N-terminal amino acid was blocked.
J Steroid Biochem Mol Biol 1995 Sep
PMID:Direct expression of pig testicular 3 alpha/beta (20 beta)-hydroxysteroid dehydrogenase in Escherichia coli. 757 8

Several bands of hydridization are detected when southern blots of human genomic DNA are proved with cDNA of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) type I. Two experimental approaches were adopted to estimate the size of the 3 beta-HSD gene family. Firstly, primer designed to amplify 3 beta-HSD type I and II genes were found on occasion to amplify DNA products of appropriate length but which were resolved as distinct sequences by denaturing gradient gel electrophoresis (DGGE). Five of these novel bands were cloned and their sequences were found to be closely related to 3 beta-HSD types I and II. Secondly, 57 genomic clones were selected from two lambda genomic libraries by hybridization with exonic probes of 3 beta -HSD type I. These were screened for novel members of the gene family by pcr amplification using various combinations of PCR primers to the type I and II genes, particularly those primers that previously amplified novel PCR products from genomic DNA. Amplification products from (lambda) clones were screened for novel sequences by DGGE. As a result of these approaches, at least five new members of the 3 beta-HSD gene family were found, one of which locates to the 3 beta -HSD type I and II gene cluster on 1p13. The existence of additional closely related but distinct members of the gene family should be recognized as a potential complication when screening PCR fragments for mutations in the type I and II genes. DGGE was found to be an exceedingly rapid means of screening amplification products from (lambda) clones to search for novel members of the gene family.
Mol Cell Probes 1995 Apr
PMID:New members of the 3 beta-hydroxysteroid dehydrogenase gene family. 760 71

The classical form of the enzyme 5-ene-3 beta-hydroxysteroid dehydrogenase/isomerase (3 beta HSD), expressed in adrenal glands and gonads, catalyzes the conversion of 5-ene-3 beta-hydroxysteroids to 4-ene-3-ketosteroids, an essential step in the biosynthesis of all active steroid hormones. To date, four distinct mouse 3 beta HSD cDNAs have been isolated and characterized. These cDNAs are expressed in a tissue-specific manner and encode proteins of two functional classes. Mouse 3 beta HSD I and III function as 3 beta-hydroxysteroid dehydrogenases and 5-en-->4-en isomerases using NAD+ as a cofactor. The enzymatic function of 3 beta HSD II has not been completely characterized. Mouse 3 beta HSD IV functions only as a 3-ketosteroid reductase using NADPH as a cofactor. The predicted amino acid sequences of the four isoforms exhibit a high degree of identity. Forms II and III are 85 and 83% homologous to form I. Form IV is most distant from the other three with 77 and 73% sequence identity to I and III, respectively. 3 beta HSD I is expressed in the gonads and adrenal glands of the adult mouse. 3 beta HSD II and III are expressed in the kidney and liver with the expression of form II greater in kidney and form III greater in liver. Form IV is expressed exclusively in the kidney. Although the amino acid composition of forms I, III and IV predicts proteins of the same molecular weight, the proteins have different mobilities on SDS-polyacrylamide gel electrophoresis. This characteristic allows for differential identification of the expressed proteins. The four structural genes encoding the different isoforms are closely linked within a segment of mouse chromosome 3 that is conserved on human chromosome 1.
J Steroid Biochem Mol Biol 1995 Jun
PMID:The murine 3 beta-hydroxysteroid dehydrogenase multigene family: structure, function and tissue-specific expression. 762 43

In the brain, the 5 alpha-reductase converting testosterone (T) is present both in neurons and in glial cells, even if it prevails in neurons; the 3 alpha-hydroxysteroid-dehydrogenase (3 alpha-HSD), the enzyme converting dihydrotestosterone (DHT) into 3 alpha-diol, is particularly concentrated in type 1 astrocytes. In glial cells, since the 5 alpha-reductase is activated by a cAMP analogue, PKA seems to be involved in the control of this enzyme, postulating that nervous inputs utilizing cAMP as the second messenger might modify the activity of this enzyme in glial cells. Moreover, the results indicate that, in type 1 astrocytes, both the 5 alpha-reductase and the 3 alpha-HSD are stimulated by the co-culture with neurons and by the addition of neuron-conditioned medium, suggesting that secretory products released by neurons might intervene in the control of glial cell function.
J Steroid Biochem Mol Biol 1995 Jun
PMID:Metabolism of steroids in pure cultures of neurons and glial cells: role of intracellular signalling. 762 76

The enzyme 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) catalyzes the 17 beta-oxidation/reduction of C18- and C19-steroids in a variety of tissues. Three human genes encoding isozymes of 17 beta-HSD, designated 17 beta-HSD types 1, 2 and 3 have been cloned. 17 beta-HSD type 1 (also referred to as estradiol 17 beta-dehydrogenase) catalyzes the conversion of estrone to estradiol, primarily in the ovary and placenta. The 17 beta-HSD type 2 is expressed to high levels in the liver, secretory endometrium and placenta. The type 2 isozyme catalyzes the oxidation of androgens and estrogens equally efficiently. Also, the enzyme possesses 20 alpha-HSD activity demonstrated by its ability to convert 20 alpha-dihydroprogesterone to progesterone. Testicular 17 beta-HSD type 3 catalyzes the conversion of androstenedione to testosterone, dehydroepiandrosterone to 5-androstenediol and estrone to estradiol. The 17 beta-HSD3 gene is mutated in male pseudohermaphrodites with the genetic disease 17 beta-HSD deficiency.
J Steroid Biochem Mol Biol 1995 Jun
PMID:The molecular biology of androgenic 17 beta-hydroxysteroid dehydrogenases. 762 83

We have previously described two distinct isoforms of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) with respect to enzymatic activity in the ovine liver and kidney. To determine which isoform(s) is expressed in the ovine placenta, we studied the characteristics of 11 beta-HSD activity in placental tissues collected at days 140-143 of pregnancy. 11 beta-HSD activity was determined by a radiometric conversion assay using cortisol and cortisone as physiological substrates. At 100 nM cortisol, the placental 11 beta-HSD utilized NAD as cofactor, but displayed preference for NADP at 10 microM cortisol. Kinetic characteristics were examined in the presence of alternate cofactors, in order to determine whether this difference in the cofactor requirement represents distinct enzymes. With NAD as cofactor, the placental 11 beta-dehydrogenase had a Km (110 +/- 18 nM) compatible with the kidney enzyme, but displayed a Km (12 +/- 2 microM) similar/identical to the liver 11 beta-HSD when NADP was used. By contrast, the placental 11-oxoreductase showed preference for NADPH regardless of cortisone concentration. Kinetic analysis, using NADPH as cofactor, revealed a single species of 11-oxoreductase activity with a Km of 4 +/- 0.9 microM and a Vmax of 3.1 +/- 0.5 pmol/mg/min. Finally, since the NAD-dependent 11 beta-HSD in the ovine placenta displayed similar/identical kinetic characteristics to the enzyme described previously in the ovine kidney where a truncated 11 beta-HSD transcript was identified, we have also determined whether this transcript is expressed in the placenta by Northern blotting. It was found that the truncated 11 beta-HSD transcript was undetectable in the total RNA samples. These results demonstrate that both liver- and kidney-types of 11 beta-HSD activities are expressed in the ovine placenta, thus providing further evidence for the existence of a NAD-dependent 11 beta-HSD distinct from the well-characterized hepatic NADP-dependent enzyme. Furthermore, the lack of the truncated 11 beta-HSD transcript in the placenta suggests that the NAD-dependent enzyme identified in placenta and kidney is the product of a gene distinct from 11 beta-HSD.
J Steroid Biochem Mol Biol 1995 Apr
PMID:Co-expression of two distinct isoforms of 11 beta-hydroxysteroid dehydrogenase in the ovine placenta. 773 1


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