Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01189 (beta-endorphin)
 21,003 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adrenal mitochondrial cytochrome P-450 which functions in cholesterol side chain cleavage (P-450scc) exhibited type I (lambdamax 385, lambdamin 420 nm) and inverse type I (lambdamin 385, lambdamax 420 nm) difference spectra with several steroids. The magnitude and type of response were dependent on the particular steroid and on the extent to which cholesterol was bound to the cytochrome in the intact mitochondrion. the inverse type I difference spectrum induced by 3beta-hydroxy-pregn-5-ene-20-one (pregnenolone) was dependent on the proportion of high spin cholesterol-cytochrome P-450scc complexes. With rat adrenal mitochondria cholest-5-ene-3beta, 20alpha-diol (20alpha-hydroxycholesterol) invariably induced a smaller inverse type I response and, under conditions where cytochrome P-450scc was nearly free of cholesterol, even produced a small type I response. Two distinct steroid binding sites on cytochrome P-450scc were detected by, respectively, the slow type I response to cholest-5-ene-3beta, 25-diol (25-hydroxycholesterol) and the rapid type I response to a subsequent addition of cholest-5-ene-3beta, 20alpha, 22 R-triol (20alpha, 22R-dihydroxycholesterol). The relative proportions of the spectral responses to these steroids were dependent on the previous extent of adrenal activation by adrenocorticotropic hormone (ACTH), because this stimulatory process altered the combination of mitochondrial cholesterol with cytochrome P-450scc. It is proposed that the two steroid binding sites on cytochrome P-450scc interact with steroids in the following way: site I binds cholesterol, 25-hydroxycholesterol, and 20alpha, 22R-dihydroxycholesterol with formation of a partially high spin cytochrome; site II binds both pregnenolone and 20alpha-OH cholesterol resulting in a low spin cytochrome. Interactions between sites I and II are not competitive, and occupancy of site II ensures a low spin state irrespective of the occupancy of site I. A second mode of interaction by 20alpha, 22R-dihydroxycholesterol stabilizes a high spin cytochrome and is competitive with site II binding by 20alpha-hydroxycholesterol or pregnenolone. Formation of a maximally high spin cytochrome follows occupancy by 20alpha, 22R-dihydroxycholesterol at both sites.
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PMID:Cytochrome P-450 of adrenal mitochondria. Spin states as detected by difference spectroscopy. 16

Steroid-induced difference spectra have been used to examine the combination of cholesterol with adrenal mitochondrial cytochrome P-450 which participates in cholesterol side chain cleavage (P-450scc) and the depletion of cholesterol from the cytochrome which results from turnover of the enzyme system. Type I difference spectra-induced by cholest-5-ene-3beta, 25-diol (25-hydroxycholesterol) and cholest-5-ene-3beta, 20 alpha, 22R-triol (20alpha, 22R dihydroxycholesterol) have been used to quantitate binding of cholesterol to two sites (I and II) on cytochrome P-450scc. The action of adrenocorticotropic hormone (ACTH) in vivo and the action of calcium or phosphate ions on isolated mitochondria stimulate the combination of cholesterol with site I but not site II. Cholesterol derived from lecithin-cholesterol micelles, however, binds to both sites. Malate-induced cholesterol depletion occurred at a comparable rate to the transfer of cholesterol from lecithin-cholesterol micelles. However, a residual proportion of cholesterol-cytochrome P-450scc complexes remained, even after 10 min of exposure to malate, and was of similar magnitude in mitochondria from both cycloheximide-treated and stressed rats. It is suggested that this reflects a less reactive form of cholesterol-cytochrome complex. Steroid-induced difference spectra indicate that sites I and II on cytochrome P-450scc are similarly depleted after metabolism of mitochondrial cholesterol in vitro and after inhibition of the action of ACTH in vivo. Anaerobiosis of adrenal cells after excision of the accumulation of cholesterol at cytochrome P-450cc. When anaerobiosis was prevented, cytochrome P-450scc in the freshly isolated mitochondria was apparently essentially free of complexed cholesterol, irrespective of the extent of ACTH action. For 30 min after suspension of the mitochondria in 0.25 M sucrose at 4 degrees, cholesterol combines with cytochrome P-450scc. The extent of this process was not affected by the presence of cycloheximide during ether stress treatment of the rats. It is concluded that there are at least two pools of mitochondrial cholesterol with access to cytochrome P-450scc but that ACTH stimulates only the pool which most readily interacts with the cytochrome.
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PMID:Cytochrome P-450 of adrenal mitochondria. In vitro and in vivo changes in spin states. 16 1

Low and high spin ferric cytochrome P-450 and reduced adrenal ferredoxin (adrenodoxin) have been directly studied by EPR techniques in whole rat adrenal glands. The spectra obtained correspond closely to those obtained from sub-cellular fractions except in the case of low spin ferric cytochrome P-450, where there are differences in the shape of the g = 2.41 line. The relative magnitudes of these peaks in anaerobic and aerobic rapidly frozen adrenals from control and corticotropin stimulated hypophysectomised rats were used to investigate the control and rate limiting steps in adrenal steroid biosynthesis via cytochrome P-450. All adrenals showed a close to maximal level of reduced adrenodoxin and aerobic and anaerobic glands from control rats and aerobic glands from corticotropin stimulated rats showed similar quantities of low spin ferric cytochrome P-450. On anaerobiosis the quantity of low spin ferric cytochrome in adrenals from corticotropin stimulated rats dropped to 30--40% of the aerobic level. Treatment of the rats with cycloheximide prior to administration of corticotropin prevented these changes. Approximately 0.4% of the total cytochrome P-450 was high spin ferric in control adrenals and in aerobic stimulated adrenals this rose to approximately to 0.6%. These results demonstrate that association of substrate with cytochrome P-450 is the rate limiting step in adrenal steroidogenesis via cytochrome P-450. It is suggested on the basis of these and mitochondrial optical and EPR experiments that the limiting step being observed is cholesterol binding to cholesterol side chain cleavage cytochrome P-450, and that the rate of this association is stimulated by corticotropin.
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PMID:Electron paramagnetic resonance studies of cytochrome P-450 and adrenal ferredoxin in single whole rat adrenal glands. Effect of corticotropin. 18 43

Steroidogenesis by adrenal mitochondria from endogenous precursors is stimulated by corticotropin (ACTH) and is sensitive to the protein-synthesis inhibitor cycloheximide. In the present investigation the effect of cycloheximide treatment on the metabolism of a number of analogues of the normal steroidogenic substrate, i.e. cholesterol, by rat adrenal mitochondria was studied. It was observed that the metabolism of analogues such as desmosterol, 26-norcholest-5-en-3beta-ol and 5-cholen-3beta-ol (that is with non-polar alkyl side chains like cholesterol), was sensitive to cycloheximide treatment. By contrast, the metabolism of those analogues with polar groupings on the side chain, i.e., 20alpha-, 24-, 25- and 26-hydroxycholesterols was insensitive to pretreatment with cycloheximide. The binding of added sterol to the cytochrome P-450 component of the mitochondrial sterol desmolase was studied. Similar studies on the equilibration time on addition of exogenous sterols to achieve maximum rates of pregnenolone production were also made. Both studies show that cholesterol, a non-polar sterol, penetrated slowly through the mitochondrial milieu to reach the cytochrome P-450 reaction centre whereas 24- and 26-hydroxycholesterols rapidly attained the enzymic environment. The cycloheximide-sensitive process in sterol metabolism appeared related to the transfer of non-polar sterols such as cholesterol within the mitochondria to a region in close proximity to the enzyme. The importance, and possible mechanism of action, of the cycloheximide-sensitive factor in the control of adrenal steroidogenesis is discussed.
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PMID:Control of sterol metabolism in rat adrenal mitochondria. 72 74

Spironolactone administration (50 mg/kg/day for 3 days) to make guinea pigs decreased cortisol production by adrenal slices in vitro. Adrenal microsomal and mitochondrial cytochrome P-450 levels were also decreased after treatment with spironolactone. The decline in adrenal cytochrome P-450 content was accompanied by decreases in microsomal 21-hydroxylase and mitochondrial cholesterol side-chain cleavage and 11beta-hydroxylase activities. Activities of other adrenal enzymes, such as delta4-hydrogenase and NADPH-cytochrome c reductase, were unaffected by spironolactone treatment. Cortisone administration to guinea pigs failed to mimic the effects of spironolactone on adrenal function, which indicates specificity of spironolactone action and excludes inhibition of adrenocorticotropin secretion as a mode of action. Addition of spironolactone to isolated adrenal mitochondria or microsomes produced type I spectral changes with spectral dissociation constants similar to those for endogenous steroid substrates. Spironolactone, in vitro, inhibited 11beta- but not 21-hydroxylase activity. The results indicate that spironolactone administration diminishes the activity of adrenal mitochondrial as well as microsomal cytochrome P-450-containing enzymes, resulting in a fall in corticosteroid output.
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PMID:Mechanism of action of spironolactone on adrenocortical function in guinea pigs. 97 70

The iron-containing protein cytochrome P-450 is present in high concentration in the adrenal cortex and is involved in the synthesis of corticosterone. This study was designed to determine the cortisol response to adrenocorticotropin (ACTH) in patients with severe iron deficiency. Eleven patients with iron deficiency and 15 normal controls were studied. Fasting blood samples were taken from all the subjects before and 30, 60 and 120 min after infusion of 25 units of ACTH for plasma cortisol determination. Six patients had blood samples collected at night, too. The same test was performed in 6 patients with iron deficiency, 7 days after therapy with 800 mg of ferrous sulfate. No significant differences were observed between patients and controls for the baseline cortisol values. The cortisol secretion and the increment at 30, 60 and 120 min after ACTH infusion were significantly lower in patients than in controls, either before or after ferrous sulfate therapy. There were no significant differences between baseline and stimulated cortisol values in patients before and after 7 days of ferrous sulfate therapy. There was no change in cortisol secretion rhythm in patients with iron deficiency (cortisol level at night = 5.1 +/- 4.3 micrograms/dl). In conclusion, the results of the present study showed that, in patients with severe iron deficiency, the cortisol secretion after ACTH stimulation was decreased.
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PMID:Reduced cortisol secretion in patients with iron deficiency. 165 78

Transforming growth factor beta 1 (TGF-beta 1) has been shown previously to induce striking alterations of bovine adrenocortical cell steroidogenic functions. One of the major lesions was characterized as a loss of steroid-17 alpha-hydroxylase activity, a key step in the biosynthetic pathway leading to active corticosteroid hormones. The mechanism of this negative effect of TGF-beta 1 on adrenocortical differentiated functions was investigated. It was observed that: 1) bovine adrenocortical 17 alpha-hydroxylase activity rapidly decreased in cells exposed to TGF-beta 1, in a time (10-20 h)-dependent manner; 2) immunoblotting of the corresponding cytochrome P-450(17) alpha showed that the loss of activity was superimposable to the decrease of the cellular protein content; 3) the cell content in 17 alpha-hydroxylase messenger RNA sharply dropped under TGF-beta 1 treatment (70-75% loss within 3-4 h) as determined by Northern blot analysis; 4) TGF-beta 1 inhibited as well the induction of P-450(17) alpha normally observed under adrenocorticotropin treatment; and 5) these TGF-beta 1 effects were selectively directed toward P-450(17) alpha expression, whereas another major steroidogenic cytochrome, i.e. P-450scc, was not affected. These observations showed that TGF-beta 1 is a potent negative modulator of 17 alpha-hydroxylase expression in bovine adrenocortical cells, very possibly at the transcriptional level. TGF-beta 1 (whose gene is expressed in these cells) may thus be examined as a possible autocrine inhibitory factor implied in the regulation of adrenocortical differentiated functions, in balance with ACTH, which represents the major positive signal in this system.
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PMID:Transforming growth factor beta 1 is a negative regulator of steroid 17 alpha-hydroxylase expression in bovine adrenocortical cells. 198 28

We have developed a method that separates rat adrenocortical cells by density into populations which retain zone specific properties in primary culture. Two different parenchymal populations were obtained and designated 2FASC (1.034 g/ml, 18.0 microns cell diameter) and 7GLOM (1.069 g/ml, 11.7 microns cell diameter). In freshly isolated cell suspensions the physical characteristics and differential steroidogenic responses to adrenocorticotropin and angiotensin II suggested that 2FASC cells originated predominantly from the zona fasciculata and 7GLOM cells from the zona glomerulosa. In primary culture (Dulbecco's Modified Eagle's Medium-F12 medium with 15% horse serum and 2.5% fetal bovine serum) the two populations exhibited different morphologies. 2FASC cells retained lipid and formed cohesive epithelial monolayers that remained stationary for 3 wk. 7GLOM cells were initially epithelial but rapidly lost lipid, spread, and assumed fibroblastic shapes. Both cell types were positive for the cholesterol side-chain cleavage cytochrome P-450 by immunofluorescence. Therefore, the morphologic changes seen in 7GLOM cultures were due to modulation, not fibroblastic overgrowth. This phenotypic plasticity may reflect the mesodermal origin of the adrenal cortex, and the subcapsular location of 7GLOM cells in vivo. In contrast, cells such as 2FASC which are located deeper in the cortex seem to have a more restricted, fully committed parenchymal phenotype.
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PMID:Density separation of rat adrenocortical cells: morphology, steroidogenesis, and P-450scc expression in primary culture. 216 28

A recombinant cDNA clone, PBC21-1, specific for bovine steroid 21-hydroxylase cytochrome P-450 (P-450C21) was identified in a bovine adrenocortical cDNA library, and this identity was confirmed by nucleotide sequencing which revealed significant amino acid homology (77%) with human P-450C21 cDNA. The pBC21-1 insert is 1.7 kilobases in length and includes a 1128 base pair region that encodes the C-terminal 376 amino acids of bovine P-450C21 as well as 535 base pairs of 3'-untranslated sequence. A novel feature of this insert is a 20 base pair intervening sequence near the 5' end, apparently the result of an aberrant splicing event. Northern blot analysis reveals that bovine P-450C21 is encoded by two transcripts, 2.3 and 2.0 kilobases in length which are detected in adrenal cortical RNA. Bovine liver, heart, kidney, and corpus luteum do not contain detectable P-450C21 transcripts. Regulation of P-450C21 gene expression by adrenocorticotropin was investigated with pBC21-1 and bovine adrenocortical cells in primary, monolayer culture. Treatment with ACTH or analogues of cAMP increases the steady-state levels of P-450C21 RNA in such cell cultures. In vitro transcription run-on assays suggest that this increase is, at least in part, due to the enhanced transcriptional activity of the P-450C21 gene.
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PMID:Bovine steroid 21-hydroxylase: regulation of biosynthesis. 242 92

The effects of adrenocorticotropin (ACTH) on 17 alpha-hydroxylase activity and cytochrome P-450(17 alpha) synthesis have been studied utilizing bovine adrenocortical cells in primary monolayer culture. A 20-fold stimulation of the conversion of pregnenolone to 17 alpha-hydroxypregnenolone was observed in postmitochondrial supernatant fractions from cells treated with ACTH as compared to controls. This increase in 17 alpha-hydroxylase activity was found to be due to a change in the Vmax and not a change in the Km(app). By immunoisolation of newly synthesized protein from an RNA-directed cell-free translation system we found that the level of P-450(17 alpha) was many-fold greater when RNA from ACTH-stimulated cells was used, as compared to RNA from control cells. A similar pattern was obtained when the rate of P-450(17 alpha) synthesis was analyzed by immunoisolation from radiolabeled cellular protein from ACTH-stimulated cells. When the total amount of P-450(17 alpha) was measured by immunoblotting we found that the levels of enzyme present correlated with the 17 alpha-hydroxylase activity. In addition, we found that these ACTH-mediated effects could be mimicked by treatment of cells with analogs of cyclic AMP. These results indicate that the activity of P-450(17 alpha) is regulated primarily by cyclic AMP-mediated changes in synthesis, probably at the transcriptional level, which in turn has a profound effect on the pattern of steroid secretion. Thus, we believe cytochrome P-450(17 alpha) to be a key regulatory enzyme in the steroidogenic pathway.
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PMID:Effects of adrenocorticotropin on 17 alpha-hydroxylase activity and cytochrome P-450(17 alpha) synthesis in bovine adrenocortical cells. 298 74


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