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Pregnenolone

About: Pregnenolone is a research topic. Over the lifetime, 3539 publications have been published within this topic receiving 126444 citations. The topic is also known as: (3b)-3-hydroxy-Pregn-5-en-20-one & 3-Hydroxypregn-5-en-20-one.


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Journal ArticleDOI
TL;DR: The quantitatively important metabolism to 7 alpha-OH compounds may contribute to the control of neurosteroid activity in brain, and derivatives of DHEA and PREG are known to be neuroactive.
Abstract: Two 'neurosteroids', dehydroepiandrosterone (DHEA) and pregnenolone (PREG), are converted by rat brain microsomes into polar metabolites, identified as the respective 7 alpha-hydroxylated (7 alpha-OH) derivatives by the 'twin ion' technique of glc-ms with deuterated substrates The enzymic reaction requires NADPH and is stimulated 2-4-fold by EDTA Under optimal conditions (pH 74, 05 mM-NADPH, 1 mM-EDTA), the Km values for DHEA and PREG are 138 and 44 microM respectively, and the Vmax values are 322 and 388 pmol/min per mg of microsomal protein respectively Trace amounts of putative 7 beta-OH derivatives of DHEA and PREG are detected Oestradiol, at a pharmacological concentration of 5 microM, inhibits DHEA and PREG 7 alpha-hydroxylation Formation of 7 alpha-hydroxylated metabolites is low in prepubertal rats and increases 5-fold in adults Derivatives of PREG and DHEA, such as PREG sulphate, DHEA sulphate, progesterone and 3 alpha-hydroxy-5 alpha-pregnan-20-one, are known to be neuroactive Therefore the quantitatively important metabolism to 7 alpha-OH compounds may contribute to the control of neurosteroid activity in brain

166 citations

Journal ArticleDOI
TL;DR: The data offer the first evidence that R347 is a crucial component of the site at which b5 interacts with the P450c17 x OR complex to promote electron transfer, and prove that the lyase activity was disrupted by interfering with electron transfer.
Abstract: Cytochrome P450c17 catalyzes steroid 17alpha-hydroxylase and 17,20-lyase activities and hence is a key enzyme in the production of human glucocorticoids and sex steroids. These two activities are catalyzed in a single substrate-binding site but are regulated independently in human physiology. We have recently shown that cytochrome b5 facilitates 17,20-lyase activity by allosterically promoting the interaction of P450c17 with P450 oxidoreductase (OR) and that the human P450c17 mutations, R347H and R358Q, selectively destroy 17,20-lyase activity while sparing 17alpha-hydroxylase activity. We transfected COS-1 cells with vectors for these P450c17 mutants and found that an excess of OR and b5 restored a small amount of 17,20-lyase activity, suggesting the mutations interfere with electron donation. To determine whether these mutations selectively interfere with the interaction of P450c17 and its electron-donating system, we expressed each P450cl7 mutant in yeast with or without OR, b5, or both, and measured enzyme kinetics in yeast microsomes using pregnenolone and 17alpha-hydroxypregnenolone as substrates. The apparent Michaelis-Menten (Km) values for the R347H mutant with and without coexpressed OR were 0.2 and 0.6 microM, respectively, and for the R358Q mutant with and without OR they were 0.3 and 0.4 microM, respectively; these values did not differ significantly from the wild-type values of 0.4 and 0.8 microM with and without OR, respectively. Furthermore, coincubation with 17alpha-hydroxypregnenolone showed a competitive mechanism for interference of catalysis. The similar kinetics and the competitive inhibition prove that the mutations did not affect the active site. Coexpression of the mutants with OR yielded insignificant 17,20-lyase activity, but addition of a 30:1 molar excess cytochrome b5 to these microsomes restored partial 17,20-lyase activity, with the R358Q mutant achieving twice the activity of the R347H mutant. These data indicate that both mutations selectively interfere with 17,20-lyase activity by altering the interaction of P450c17 with OR, thus proving that the lyase activity was disrupted by interfering with electron transfer. Furthermore, the data offer the first evidence that R347 is a crucial component of the site at which b5 interacts with the P450c17 x OR complex to promote electron transfer.

165 citations

Journal ArticleDOI
TL;DR: The two-hit model, which considers the persistence of StAR-independent steroidogenesis and the differences in the fetal and postnatal ages at which the testis, adrenal zona glomerulosa, adrenAL zona fasciculata and ovary are stimulated, predicts and explains all of the various clinical manifestations of lipoid CAH.

163 citations

Journal ArticleDOI
TL;DR: The often confusing literature on StAR’s mechanism of action is examined, particularly in light of recent work establishing the importance of other players, and a model for StAR's interaction with cholesterol and with some of these other proteins is presented.
Abstract: The adrenal cortex is a tissue of excess in terms of both cholesterol metabolism and cholesterol exchange with the circulation. Exceptionally high levels of lipoprotein receptors in this highly vascularized tissue provide ready access to dietary cholesterol, allowing the adrenocortical cells to maintain impressive stores of cytoplasmic cholesterol ester (CE) droplets. Tightly packed among the CE droplets are specialized mitochondria, carrying in their inner membranes high levels of the cytochrome P450scc (CYP11A1). This enzyme carries out the so-called side chain cleavage reaction, consuming cholesterol to produce pregnenolone, the precursor of cortisol and all other steroids. Glucocorticoid synthesis is tightly regulated at the level of cholesterol metabolism, which responds to ACTH stimulation over a period of minutes and ceases equally quickly when this hormone is removed. Remarkably, this dynamic process is modulated under most circumstances not by control of the intrinsic enzymatic activity of P450scc, but rather by substrate availability. For this reason, cholesterol transport within the mitochondrion has emerged as the key control point for steroidogenesis. The adrenal cortex is not alone in requiring efficient and controlled delivery of cholesterol into mitochondria. Other steroidogenic cells, including several cell types in the ovary, the Leydig cells of the testis, and a subset of hippocampal neurons (1), also employ P450scc to produce pregnenolone and a variety of downstream steroid hormones or neurosteroids. In vertebrates ranging from birds and fish (2) to mammals, these various cell types all express a short-lived mitochondrial import factor now called the steroidogenic acute regulatory protein (StAR), which mediates this process. Here, I examine the often confusing literature on StAR’s mechanism of action, particularly in light of recent work establishing the importance of other players, and I present a model for StAR’s interaction with cholesterol and with some of these other proteins. I also discuss the insights into mitochondrial function that have come from the analysis of patients with congenital adrenal hyperplasia (CAH), who lack this factor. Finally, I consider the multi-tiered regulation of StAR and related proteins in adrenocortical cells and other steroidogenic cell types.

163 citations

Journal ArticleDOI
TL;DR: Findings demonstrate that residues on the NR2 subunit are key determinants of modulation by PS and 3α5βS, and indicate that it may be possible to develop therapeutic agents that target steroid modulatory sites of specific NMDA receptor subtypes.
Abstract: 1 The neurosteroid pregnenolone sulphate (PS) potentiates N-methyl-D-aspartate (NMDA) receptor mediated responses in various neuronal preparations. The NR1 subunit can combine with NR2A, NR2B, NR2C, or NR2D subunits to form functional receptors. DiAerential NR2 subunit expression in brain and during development raises the question of how the NR2 subunit influences NMDA receptor modulation by neuroactive steroids. 2 We examined the eAects of PS on the four diheteromeric NMDA receptor subtypes generated by co-expressing the NR1100 subunit with each of the four NR2 subunits in Xenopus oocytes. Whereas PS potentiated NMDA-, glutamate-, and glycine-induced currents of NR1/NR2A and NR1/NR2B receptors, it was inhibitory at NR1/NR2C and NR1/NR2D receptors. 3 In contrast, pregnanolone sulphate (3a5bS), a negative modulator of the NMDA receptor that acts at a distinct site from PS, inhibited all four subtypes, but was approximately 4 fold more potent at NR1/NR2C and NR1/NR2D than at NR1/NR2A and NR1/NR2B receptors. 4 These findings demonstrate that residues on the NR2 subunit are key determinants of modulation by PS and 3a5bS. The modulatory eAects of PS, but not 3a5bS, on dose-response curves for NMDA, glutamate, and glycine are consistent with a two-state model in which PS either stabilizes or destabilizes the active state of the receptor, depending upon which NR2 subunit is present. 5 The selectivity of sulphated steroid modulators for NMDA receptors of specific subunit composition is consistent with a neuromodulatory role for endogenous sulphated steroids. The results indicate that it may be possible to develop therapeutic agents that target steroid modulatory sites of specific NMDA receptor subtypes. British Journal of Pharmacology (2002) 135, 901‐909

162 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
202344
202255
202124
202028
201950
201835