Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.1.1.3 (HSD)
3,464 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The apoenzyme and holoenzyme (NADP+ complex) of human placental 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) were prepared from affinity chromatography using various elutions by column liquid chromatography. The apoenzyme was obtained using NAD+ elution in a Blue-Sepharose column, followed by NAD+ separation on a Phenyl-Superose hydrophobic-interaction or a Mono Q anion-exchange column. The 17 beta-HSD-NADP+ complex was prepared using NADP+ elution in a Blue-Sepharose column. The two forms have different A280/A260 ratios and are suitable for further study of enzyme-cofactor interactions.
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PMID:Preparation of the apoenzyme and holoenzyme forms of human 17 beta-hydroxysteroid dehydrogenase. 849 77

Excess glucocorticoids impair fetal growth and cause teratogenesis. Placental 11 beta-hydroxysteroid dehydrogenase (11 beta HSD) catalyzes the inactivation of cortisol to cortisone, preventing the high maternal cortisol levels from reaching the fetal circulation and thus preserving the low cortisol fetal environment. In previous work, an NADP-dependent isoform of 11 beta HSD has been purified from rat liver, a cDNA isolated, and the human homolog cloned. However, much evidence suggests tissue-specific 11 beta HSD activities that cannot be explained by the liver-type isoform. Therefore, we have partially purified human placental 11 beta HSD and compared it to the enzyme in rat liver. Human placental subcellular fractions exhibited NAD-dependent 11 beta HSD activity, but showed little activity with NADP. The enzyme had a pH optimum of 7-8.5 (peak, 7.7), was only sparingly soluble in detergents (solubility with Triton X-100 was very poor), and exhibited little latency or change in pH profile in detergent solution. By contrast, rat liver 11 beta HSD was exclusively NADP dependent and was easily solubilized by a wide range of detergents (including Triton X-100), revealing substantial latency and altered pH profile [optimum of 10, becoming 7-10 (peak, 9.5) in detergent]. These data do not merely reflect species differences, as rat placental 11 beta HSD was similar to the human placental isoform. AMP affinity chromatography, which was completely without affinity for rat liver 11 beta HSD, achieved a 1000-fold purification of human placental 11 beta HSD. This had Km values for corticosterone (mean +/- SE, 14 +/- 1 nM) and cortisol (approximately 55 nM) that were over 100 times lower than that for liver 11 beta HSD. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis allowed identification of a band (apparent mol wt, 40,000) that correlated consistently with human placental 11 beta HSD activity (contrasting with a mol wt of 34,000 for rat liver 11 beta HSD). Thus, the NAD-dependent human placental 11 beta HSD is distinct from the previously characterized rat liver isoform and may be the product of a separate gene.
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PMID:Human placental 11 beta-hydroxysteroid dehydrogenase: evidence for and partial purification of a distinct NAD-dependent isoform. 850 62

11 beta-Hydroxysteroid dehydrogenase (11 beta HSD) converts glucocorticoids to inactive products and is thus thought to confer specificity for aldosterone on the type I mineralocorticoid receptor in the kidney. Recent studies indicate the presence of at least two isozymes of 11 beta HSD. In vitro, the NAD(+)-dependent kidney (type 2) isozyme catalyzes 11 beta-dehydrogenase but not reductase reactions, whereas the NADP(+)-dependent liver (type 1) isozyme catalyzes both reactions. We have now characterized the human gene encoding kidney 11 beta HSD (HSD11K). A bacteriophage P1 clone was isolated after screening a human genomic library by hybridization with sheep HSD11K cDNA. The gene consists of 5 exons spread over 6 kb. The nucleotide binding domain lies in the first and the second exon, and the catalytic domain in the fourth exon. The 5' flanking sequences and first exon are GC-rich (80%), suggesting that the gene may be transcriptionally regulated by factors that recognize GC-rich sequences. Fluorescence in situ hybridization of metaphase chromosomes with a positive P1 clone localized the gene to chromosome 16q22. In contrast, the HSD11L (liver isozyme) gene is located on chromosome 1 and contains 6 exons; the coding sequences of these genes are only 21% identical. HSD11K is expressed at high levels in the placenta and kidney of midgestation human fetuses and at lower levels in lung and testes. Different transcriptional start sites are utilized in kidney and placenta. These data should be applicable to genetic analysis of the syndrome of apparent mineralocorticoid excess, which may represent a deficiency of 11 beta HSD.
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PMID:Gene structure and chromosomal localization of the human HSD11K gene encoding the kidney (type 2) isozyme of 11 beta-hydroxysteroid dehydrogenase. 853 71

Two isoforms of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) catalyse the interconversion of active cortisol to inactive cortisone; 11 beta-HSD1 is a low affinity, NADP(H)-dependent dehydrogenase/oxo-reductase, and 11 beta-HSD2 a high affinity, NAD-dependent dehydrogenase. Because of the importance of 11 beta-HSD in regulating corticosteroid hormone action, we have analysed the distribution of the 11 beta-HSD isoforms in human adult and foetal tissues (including placenta), and, in addition have performed a series of substrate specificity studies on the novel, kidney 11 beta-HSD2 isoform. Using an RT-PCR approach, we failed to detect 11 beta-HSD1 mRNA in any human mid-gestational foetal tissues. In contrast 11 beta-HSD2 mRNA was present in foetal lung, adrenal, colon and kidney. In adult tissues 11 beta-HSD2 gene expression was confined to the mineralocorticoid target tissues, kidney and colon, whilst 11 beta-HSD1 was expressed predominantly in glucocorticoid target tissues, liver, lung, pituitary and cerebellum. In human kidney homogenates, 11-hydroxylated progesterone derivatives, glycyrrhetinic acid, corticosterone and the "end products" cortisone and 11-dehydrocorticosterone were potent inhibitors of the NAD-dependent conversion of cortisol to cortisone. Finally high levels of 11 beta-HSD2 mRNA and activity were observed in term placentae, which correlated positively with foetal weight. The tissue-specific distribution of the 11 beta-HSD isoforms is in keeping with their differential roles, 11 beta-HSD1 regulating glucocorticoid hormone action and 11 beta-HSD2 mineralocorticoid hormone action. The correlation of 11 beta-HSD2 activity in the placenta with foetal weight suggests, in addition, a crucial role for this enzyme in foetal development, possibly in mediating ontogeny of the foetal hypothalamo-pituitary-adrenal axis.
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PMID:Type 2 11 beta-hydroxysteroid dehydrogenase in foetal and adult life. 854 71

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD) catalyzes the conversion of cortisol to cortisone. This activity may be deficient in the syndrome of apparent mineralocorticoid excess (AME). 11 beta-HSD L (Type I), isolated from liver, is widely expressed and utilizes NADP+ as a cofactor. The gene for 11 beta-HSD L was found to be normal in patients of AME. A second isoform, 11 beta-HSD K (Type II), isolated from kidney, is more tissue specific in expression and utilizes NAD+ as a cofactor. The cDNA clone encoding 11 beta-HSD K was isolated from sheep kidney. The cDNA is 1.8 kb in length and encodes a protein of 404 amino acid residues with a predicted M(r) 43,953. The recombinant enzyme functions as an NAD(+)-dependent 11 beta-dehydrogenase with very high affinity for steroids, but it has no detectable reductase activity. It is 37% identical in amino acid sequence to an NAD(+)-dependent isozyme of 17 beta-hydroxysteroid dehydrogenase. It is expressed at high levels in the kidney, placenta, adrenal and at lower levels in colon, stomach, heart and skin. The human 11 beta-HSD K gene consists of five exons spread over 6 kb. The nucleotide binding domain lies in the first and the second exon, and the catalytic domain in the fourth exon. The promoter for 11 beta-HSD K gene lacks a TATA box and has a high GC base content, suggesting that the gene may be transcriptionally regulated by factors that recognize GC-rich sequences. Fluorescent in situ hybridization of metaphase chromosomes with a positive bacteriophage P1 genomic 11 beta-HSD K clone localized the gene to chromosome 16q22. In contrast, the 11 beta-HSD L gene is located on chromosome 1 and contains 6 exons; the coding sequences of these genes are only 21% identical. Different transcriptional start sites are utilized in kidney and placenta.
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PMID:Analysis of the human gene encoding the kidney isozyme of 11 beta-hydroxysteroid dehydrogenase. 854 72

The isoenzymes of the 3 beta-hydroxysteroid dehydrogenase/5-ene-4-ene-isomerase (3 beta-HSD) gene family catalyse the transformation of all 5-ene-3 beta-hydroxysteroids into the corresponding 4-ene-3-keto-steroids and are responsible for the interconversion of 3 beta-hydroxy- and 3-keto-5 alpha-androstane steroids. The two human 3 beta-HSD genes and the three related pseudogenes are located on the chromosome 1p13.1 region, close to the centromeric marker D1Z5. The 3 beta-HSD isoenzymes prefer NAD+ to NADP+ as cofactor with the exception of the rat liver type III and mouse kidney type IV, which both prefer NADPH as cofactor for their specific 3-ketosteroid reductase activity due to the presence of Tyr36 in the rat type III and of Phe36 in mouse type IV enzymes instead of Asp36 found in other 3 beta-HSD isoenzymes. The rat types I and IV, bovine and guinea pig 3 beta-HSD proteins possess an intrinsic 17 beta-HSD activity specific to 5 alpha-androstane 17 beta-ol steroids, thus suggesting that such "secondary" activity is specifically responsible for controlling the bioavailability of the active androgen DHT. To elucidate the molecular basis of classical form of 3 beta-HSD deficiency, the structures of the types I and II 3 beta-HSD genes in 12 male pseudohermaphrodite 3 beta-HSD deficient patients as well as in four female patients were analyzed. The 14 different point mutations characterized were all detected in the type II 3 beta-HSD gene, which is the gene predominantly expressed in the adrenals and gonads, while no mutation was detected in the type I 3 beta-HSD gene predominantly expressed in the placenta and peripheral tissues. The mutant type II 3 beta-HSD enzymes carrying mutations detected in patients affected by the salt-losing form exhibit no detectable activity in intact transfected cells, at the exception of L108W and P186L proteins, which have some residual activity (approximately 1%). Mutations found in nonsalt-loser patients have some residual activity ranging from approximately 1 to approximately 10% compared to the wild-type enzyme. Characterization of mutant proteins provides unique information on the structure-function relationships of the 3 beta-HSD superfamily.
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PMID:Structure-function relationships and molecular genetics of the 3 beta-hydroxysteroid dehydrogenase gene family. 854 74

An overview of the application of kinetic methods to the delineation of 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) heterogeneity in mammalian tissues is presented. Early studies of 17 beta-HSD activity in animal liver and kidney subcellular fractions were suggestive of multiple forms of the enzyme. Subsequently, detailed characterization of activity in cytosol and subcellular membrane fractions of human placenta, with particular emphasis on inhibition kinetics, yielded evidence of two kinetically-differing forms of 17 beta-HSD in that organ. Gene cloning and transfection experiments have confirmed the identity of these two proteins as products of separate genes. 17 beta-HSD type 1 is a cytosolic enzyme highly specific for C18 steroids such as 17 beta-estradiol (E2) and estrone (E1). 17 beta-HSD type 2 is a membrane bound enzyme reactive with testosterone (T) and androstenedione (A), as well as E2 and E1. Useful parameters for the detection of multiple forms of 17 beta-HSD appear to be the E2/T activity ratio, NAD/NADP activity ratios, steroid inhibitor specificity and inhibition patterns over a wide range of putative inhibitor concentrations. Evaluation of these parameters for microsomes from samples of human breast tissue suggests the presence of 17 beta-HSD type 2. The 17 beta-HSD enzymology of human testis microsomes appears to differ from placenta. Analysis of human ovary indicates granulosa cells are particularly enriched in the type 1 enzyme with type 2-like activity in stroma/theca. Mouse ovary appears to contain forms of 17 beta-HSD which differ from 17 beta-HSD type 1 and type 2 in their kinetic properties.
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PMID:Kinetic analysis of enzymic activities: prediction of multiple forms of 17 beta-hydroxysteroid dehydrogenase. 854 76

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD) modulates glucocorticoid interactions with mineralocorticoid and glucocorticoid receptors in vivo, by converting 11 beta-hydroxyglucocorticoids to their inactive 11-ketone derivatives. Defective 11 beta-oxidation of glucocorticoids has been associated with hypertension. The objective of this study was to investigate whether 11 beta-HSD contributes to the occurrence of hypertension in spontaneously hypertensive rats (SHRs). The liver and kidney microsomal oxidations of corticosterone (the physiological glucocorticoid in rats) in organs from juvenile (3 weeks old) and adult (3 months old) SHR and Wistar-Kyoto (WKY) rats, with NAD and NADP, show no differences between rat strains. For cortisol, with NADP, adult SHRs show (1.3-3 times; P < 0.05) lower kidney microsomal oxidation rates. The liver microsomal reduction of cortisone shows remarkable interstrain differences; with NADH, reduction is conducted only by adult WKY rats, whereas with NADPH, juvenile animals show similar reduction rates, but at adulthood, only WKYs reduce cortisone. Using Western blot analysis with antibodies against 11 beta-HSD1, positive signals are obtained only for liver microsomes, appearing somewhat lower in SHRs for juvenile but not adult animals. Urinary corticosterone/11-dehydrocorticosterone ratios (measured in adult animals) are not different between rat strains, but are elevated after administration of corticosterone in both strains (although significant only in SHRs). The data provide no indications for exaggerated stimulation of renal corticosteroid receptors, due to modified 11 beta-HSD, in SHRs. However, the experiments suggest the existence of multiple 11 beta-HSDs, in addition to 11 beta-HSD1 and 11 beta-HSD2, some of which may be modified in SHR, but the nature and physiological role of these 11 beta-HSDs is unclear.
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PMID:Comparison of 11 beta-hydroxysteroid dehydrogenase in spontaneously hypertensive and Wistar-Kyoto rats. 858 2

This study evaluated the expression of the corticosteroid-metabolizing enzyme 11 beta-hydroxysteroid dehydrogenase (11 beta HSD) during in vitro decidualization of human endometrial stromal cells. The cultured stromal cells displayed both NADP(+)-dependent (type 1) and NAD(+)-dependent (type 2) 11 beta HSD activities under basal conditions. Although the cells did not respond to estradiol (E2) added alone, catalytic levels of both isoforms were enhanced by medroxyprogesterone acetate (MPA) and further enhanced by E2 plus MPA. Type I messenger RNA (mRNA) was undetected by Northern analysis of total RNA, but was evident as a 1.5-kilobase band in polyadenylated selected RNA from E2- plus MPA-treated cultures. Use of RT-PCR to augment the sensitivity of mRNA detection revealed the presence of type I mRNA as a faint band in the MPA-treated cultures and as an intense band in the E2- plus MPA-treated cultures. Thus, type I mRNA is present as a low abundance message in the cultured stromal cells whose steady state levels parallel progestin-enhanced enzyme activity. As the expression of several progestin-regulated decidualization markers is also augmented by E2, the results of the present study reveal a correlation between enhanced 11 beta HSD expression and the decidualization reaction. Time-course measurements indicated that elevated 11 beta HSD expression is an early event in the decidualization response, which precedes E2- plus MPA-enhanced PRL production by several days. Clear dose-response effects on both type 1 and type 2 11 beta HSD activities were obtained in cells incubated with 10(-8) mol/liter E2 added together with MPA at concentrations that approximated circulating progesterone levels from the luteal phase (10(-9) mol/liter) through pregnancy (10(-7) mol/liter). Corticosteroids are thought to exert toxic and teratogenic effects on the implanting embryo and could influence trophoblast invasion by regulating extracellular matrix turnover. Therefore, the novel finding that decidualization involves marked enhancement of the corticosteroid-metabolizing capacity of stromal cells suggests a mechanism by which decidual cells could affect the health and invasiveness of implanting trophoblastic cells.
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PMID:Expression of 11 beta-hydroxysteroid dehydrogenase during decidualization of human endometrial stromal cells. 859 7

Rat liver 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase (3 alpha-HSD) inactivates circulating steroid hormones and is involved in polycyclic aromatic hydrocarbon (PAH) carcinogenesis. It is the only HSD of known structure in the aldo-keto reductase (AKR) superfamily and may provide a paradigm for other mammalian HSDs in this family. The structure of the 3 alpha-HSD.NADP+ binary complex has been determined at 2.7 A resolution and refined to a crystallographic R-factor of 23.4% with good geometry. The model is similar to other binary complexes in the AKR superfamily in that NADP+ binds at the C-terminal end of an alpha/beta barrel. However, it is unique in that NADP+ is bound in two alternate conformations, probably because of the lack of a salt-linked "safety belt" over the pyrophosphate bridge. The structure supports a previously proposed catalytic mechanism for carbonyl reduction in which Tyr 55 is the general acid, and its effective pKa is lowered by the adjacent Lys 84. We present evidence that the structurally distinct short-chain dehydrogenase/reductase (SDR) superfamily may have convergently evolved a similar catalytic mechanism. Insight into substrate binding is offered by a crystal packing contact in which a neighboring molecule inserts a tryptophan residue (Trp 227) into an apolar cleft in 3 alpha-HSD. This cleft is proximal to the bound NADP+ cofactor and contains a surface of apolar residues (Leu 54, Trp 86, Leu 122, Phe 128, Phe 129, Leu 137, Phe 139), making it a likely candidate for the substrate-binding site. Thus, in forming this crystal contact, Trp 227 may mimic a portion of a bound steroid. In addition, we propose that a water molecule in the active site indicates the position of the hydroxyl oxygen in a 3 alpha-hydroxysteroid substrate. Knowledge of the position of this water molecule, combined with the stereochemistry of hydride transfer, suggests that the alpha face of a bound steroid will be oriented toward the side of the apolar cleft containing Trp 86.
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PMID:Structure of 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase complexed with NADP+. 871 59


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