Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P56851 (epididymal)
 11,273 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

As it was shown previoulsy by others, the membrane-bound phosphodiesterase (cyclic adenosine 3':5'-monophosphate phosphodiesterase) of rat epididymal fat cells was stimulated when intact cells were exposed to insulin. The levels of stimulation observed in the present study in the cell homogenate and microsomal fraction were approximately 2.0- to 2.5-fold and 2.5- to 3.0-fold, respectively, when the initial substrate level was 100 nM and insulin concentration was 1 to 3 nM. When the microsomal fraction was subjected to a sucrose density gradient centrifugation, most of the insulin-sensitive phosphodiesterase activity was fractionated into the "light" microsomal fraction which was rich in NADH2:potassium ferricyanide:oxidoreductase) and low in 5'-AMPase, adenylate cyclase, and insulin-binding activities. The latter three activities were mostly fractionated into the "heavy" microsomal fraction. Both basal and insulin-stimulated phosphodiesterase activities were low when cells were homogenized in the presence of N-ethylmaleimide or p-chloromercuribenzoate. The insulin-stimulated enzyme activity was also low when cells were homogenized in the presence of --SH compounds (e.g. dithiothreitol) or certain metal-chelating agents (e.g. ethylene glycol bis(beta-aminoethyl ehter)-N,N'-tetraacetate (EGTA)), or in a nitrogen atmosphere. The effect of EGTA was prevented by the addition of certain heavy metal ions but not by the addition of Ca2+ or Ca2+ plus Mg2+ ions. When cells were homogenized in the presence of certain oxidants (e.g. diamide, sodium tetrathionate, or air), a high plus-insulin activity was observed; this activity was not lowered by subsequent treatment of the enzyme with N-ethylmaleimede, EGTA, or fresh cell homogenate that was prepared in the presence of EGTA. However, the activity of an apparently oxidized enzyme could still be lowered by treatment woth dithiothreitol. A partially purified enzyme in the enzyme in the microsomal fraction was fairly stable both in basal and insulin-stimulated states (fully active after 35 days when kept at -20degrees). EGTA added to the homogenization buffer lowered the basal phosphodiesterase activity, but this effect was reversed by the addition of Ca2+ ions. EGTA also decreased the enzyme activity that was stimulated by norepinephrine. However, neither EGTA nor dithiothreitol had any effect on the activities of 5'-AMPase, NADH-dehydrogenase, and malate dehydrogenase of fat cells. The above data indicate that most of the insulin-sensitive phosphodiesterase and the so-called "cell membrane markers" are associated with different subcellular particles in the cell homogenate. In addition, the data seem to indicate that the insulin-stimulated phosphodiesterase has certain --SH groups and that the activity of the enzyme is stabilized when the --SH groups are oxidized by certain oxidants including molecular oxygen. It is suggested that the air oxidation of the enzyme is catalyzed by a trace of certain heavy metal ions and, therefore, can be blocked by a metal-chelating agent.
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PMID:Insulin-sensitive phosphodiesterase. Its localization, hormonal stimulation, and oxidative stabilization. 17 Feb 71

A protocol for the rapid purification of the glycerol dehydrogenase (glycerol: NAD+ 2-oxidoreductase, EC 1.1.1.6) from the thermophile Bacillus stearothermophilus has been developed using a combination of chromatographic techniques including affinity chromatography on a Sepharose-immobilised triazine dye (Procion red, HE3B, ICI). Substrate specificity has been examined and Km values determined. The protein has been shown to have an oligomeric Mr of approx. 180,000 and consists of four identical subunits of Mr 42,000. Exposure to chelating agents (e.g., EDTA) leads to total loss of activity; the EDTA-inactivated enzyme can be reactivated by Zn2+ and requires 1 mol equivalent of zinc per subunit for full catalytic activity. Other divalent cations such as Cd2+ and Co2+ will reactivate the apo-enzyme but yields an enzyme of lower specific activity. The enzyme binds 1 equivalent of NADH per subunit and during catalysis transfers the 4-pro-R hydride from the nicotinamide ring of the reduced-coenzyme to the substrate. Glycerol increases the dissociation constant for the interaction between NADH and Zn-metallo-glycerol dehydrogenase (ZnGDH) but has no effect on the equilibrium between NADH and metal-depleted enzyme.
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PMID:Isolation and characterisation of the glycerol dehydrogenase from Bacillus stearothermophilus. 249 67

A reliable isocratic high performance liquid chromatography (HPLC) procedure has been developed for the separation and subsequent radiometric quantitation of delta 4-3-ketosteroid-5 alpha-oxidoreductase enzyme activity. The specificity of this HPLC procedure has been confirmed through the use of authentic androgen radioisotopes, linearity of detector response, and mass spectral analysis. In conjunction with this we have also developed a simple Sep-Pak sample preparation procedure which allows the uniform complete isolation of these androgens from epididymal tissue homogenates. Utilizing these procedures we have determined the regional distribution of 5 alpha - reductase in the epididymis of the CD-1 mouse. Regional differences in enzyme activity were found between the caput-corpus and cauda regions. Enzyme activity (specific activity) was higher in the caput-corpus region (28.98 pmol 5 alpha - reduced androgens formed/h/mg protein) than the cauda region (6.20 pmol 5 alpha - reduced androgens formed/h/mg protein).
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PMID:Radiometric quantitation of delta 4-3-ketosteroid-5 alpha-oxidoreductase utilizing high performance liquid chromatography. 325 24

The epididymal epithelial ultrastructure has been described in the adult male North American opossum, Didelphis virginiana. Morphological results have suggested that absorptive activity is prominent in the proximal epididymal region by virtue of numerous microvilli, an endocytotic complex, dense granules, and multivesicular bodies in the apical cytoplasm. In contrast, the middle and distal epididymal regions exhibit ultrastructural features indicative of protein synthesis such as large invaginated euchromatic nuclei, large nucleoli, and increased amounts of granular endoplasmic reticulum. It is in the middle and distal epididymal regions where sperm head rotation and sperm pairing take place. Epididymal delta 4-3-ketosteroid-5 alpha-oxidoreductase (5 alpha-reductase) activity also has been measured. It has been found that the level of enzyme activity differs significantly (p less than 0.01) between the proximal, middle, and distal epididymal regions. Enzyme-specific activity has been found to be highest in the middle region (47.6 +/- 5.4 picomoles 5 alpha-reduced androgens formed/b/mg protein), lower in the distal region (18.3 +/- 0.7 picomoles 5 alpha-reduced androgens formed/b/mg protein), with little activity (2.4 +/- 1.2 picomoles 5 alpha-reduced androgens formed/h/mg protein) found in the proximal epididymal region. This regional distribution of enzyme activity differs markedly from that reported for eutherian mammals. Both the suggested epididymal protein synthetic and secretory activity and the level of epididymal 5 alpha-reductase activity appear to correlate regionally with the morphological changes that occur in the opossum spermatozoa as they transit the epididymis.
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PMID:Unique regional distribution of delta 4-3-ketosteroid-5 alpha-oxidoreductase and associated epididymal morphology in the marsupial, Didelphis virginiana. 367 95

We have investigated the effects of two 4-ene-steroid 5 alpha-reductase inhibitors, diethyl-4-methyl-3-oxo-4-aza-5 alpha-androstane-17 beta-carboxamide (4-MA) and (4R)-5,10-seco-19-norpregna-4, 5-diene-3,10,20-trione (SECO), on testicular and epididymal androgen biosynthesis. Kinetic analyses revealed that both compounds inhibited epididymal DHT biosynthesis. 4-MA was a competitive inhibitor of epididymal nuclear and microsomal 4-ene-steroid 5 alpha-reductases (3-oxo-5 alpha-steroid: NADP 4-ene-oxidoreductase EC 1.3.1.22) with Kiapp values of 12.8 and 15.1 nmol/l compared to the respective Kmapp values of 185 and 240 nmol/l. Values for the Vmaxapp were always within 70-130% of the control. SECO at 1.0 mumol/l, also inhibited epididymal nuclear and microsomal 4-ene-steroid-5 alpha-reductases, causing respectively 2.9 and 5.2-fold increases in Kmapp. The Vmaxapp values were unchanged. However, SECO concentrations of 5 and 25 mumol/l abolished 4-ene-steroid 5 alpha-reductase activity at all testosterone concentrations. To examine the specificity of these compounds, we investigated their effects on the enzymes that convert pregnenolone to testosterone. Rat testis microsomes converted pregnenolone to testosterone via the 4-ene-3-oxo pathway, with the major metabolites being progesterone, 17-hydroxyprogesterone, 4-androstenedione and testosterone; some 17-hydroxypregnenolone was also formed. Very small amounts of dehydroepiandrosterone (DHA) and 5-androstenediol were detected. SECO, at a concentration that completely inhibited epididymal 4-ene-steroid 5 alpha-reductase activity, did not alter the metabolic profile of pregnenolone metabolism. However, 4-MA prevented the appearance of 4-ene steroids, and large quantities of 17-hydroxypregnenolone and DHA accumulated, suggesting that inhibition of the 3 beta-hydroxysteroid: NAD(P)+ oxidoreductase (EC 1.1.1.51) and 3-oxosteroid 5-ene-4-ene-isomerase (EC 5.3.3.1) [3 beta-hydroxysteroid dehydrogenase-isomerase] was occurring. Optimal conditions for the microsomal conversion of DHA to 4-androstenedione were determined; kinetic analyses of the 3 beta-hydroxysteroid dehydrogenase-isomerase activity revealed that 4-MA inhibited this reaction non-competitively, reducing Vmaxapp values to 25% of the control. The Kiapp determined from the intercept replot, was 121 nmol/l, and the Kmapp was always between 90 and 130% of the control value. It is concluded that SECO is more specific than 4-MA in its effects on androgen biosynthesis in the testis and epididymis and that both these drugs should provide useful tools in assessments of the relative contributions of 5 alpha-reduced androgens to androgen dependent processes.
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PMID:The effects of diethyl-4-methyl-3-oxo-4-aza-5 alpha-androstane-17 beta-carboxamide (4-MA) and (4R)-5,10-SECO-19-norpregna-4,5-diene-3,10,20-trione (SECO) on androgen biosynthesis in the rat testis and epididymis. 370 62

The conversion of testosterone to 5 alpha-dihydrotestosterone, catalysed by 4-ene-steroid 5 alpha-reductase (3-oxo-5 alpha-steroid: NADP+ 4-ene-oxidoreductase EC 1.3.1.22) requires NADPH. In the present study, the role of flavins and Co-enzyme Q in this proton transfer was investigated for the first time in any male androgen target tissue. Flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) inhibited epididymal nuclear 4-ene-steroid 5 alpha-reductase activity non-competitively with respect to the substrate testosterone. However, neither the oxidized nor reduced forms of Co-enzyme Q affected the Kmapp or the Vmaxapp and the reduced form was unable to support catalytic activity in the absence of NADPH. Further investigation of the effects of flavins revealed that the inhibition was caused by an elevation of NADP+ in the incubations and that the incorporation of a NADPH generating system abolished the inhibition. Therefore, neither flavins nor Co-enzyme Q directly affected the 4-ene-steroid 5 alpha-reductase activity. Further evidence to support this conclusion was obtained when several inhibitors of electron transfer reactions failed to inhibit 4-ene-steroid 5 alpha-reductases from rat epididymides, prostate and seminal vesicles. These findings show that, in male rat androgen target tissues, the conversion of testosterone to 5 alpha-dihydrotestosterone does not require intermediates of electron transfer reactions. We propose that the reduction proceeds by the direct transfer of protons from NADPH to testosterone.
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PMID:Mechanism of 4-ene-steroid 5 alpha-reductase proton transfer in androgen target tissues. 674 43

The present paper describes some physiocochemical properties of the soluble 3 alpha-oxidoreductases in the rat testis and prostate, and comparison with rat epididymal 3 alpha-oxidoreductase, published previously (Hastings & Hansson 1979). The testicular enzyme shows properties very similar to that in the epididymis (size, stability, pH optium) except for minor differences in charge (iso-electric point). The prostatic enzyme revealed a slightly higher molecular weight, and was more sensitive to heating than those in the testis and epididymis, whereas the iso-electric point was the same as that in the testis (pI-5.25). The enzymes in all tissues exhibit very similar shapes (f/fo 1.14-1.17). The similar properties of the testicular and prostate 3 alpha-oxidoreductases to those previously reported for that in the epididymis may indicate that these enzymes represent identical peptide chains. The small differences observed in size, temperature stability and change may be due to their presence in different environments.
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PMID:Physico-chemical characterization of the soluble 3 alpha-hydroxysteroid oxidoreductase in the rat testis and prostate. 740 96

The aim of the paper has been to establish the manner, in which the lead damages spermatozoa, and what role in this process is played by epithelial cells of epididymis, responsible for maturation and survival of spermatozoa in that organ. The studies were carried out on sexually mature male rats, which were given to drink lead acetate (II), for 9 months. Studies in vitro were also performed on isolated, from epididymis, spermatozoa of rats untreated with lead, but incubated in milieu having high concentration of ions of that element. A number of research techniques were employed: morphologic examinations of testes and epididymidis, with stages of spermic epithelium and epididymal zones being taken into consideration; microscopic-electron studies of cells in epididymal duct wall and spermatozoa from duct lumen; X-ray microanalysis determining the presence and the types of elements on ultrathin specimens of epididymal cells and in spermatozoa; histochemical examinations for oxidoreductase of spermatozoa. Lead deposits were found in cells of the epididymal duct and lumen. The most of such deposits were revealed in epithelial cells, through which substances from the blood vessels are transported to the duct lumen-to spermatozoa. That gives rise to the possibility of damaging spermatozoa, which had been verified by microscopic-electron and histochemical examinations of spermatozoa. The vitro studies disclosed the ability of lead to penetrate spermatozoa, particularly the midpiece. Very important is the conclusion highlighting that the presence of lead deposits in epithelial cells and in the lumen of epididymal duct supports the possibility of excreting this element from the organism of mammals with the semen.
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PMID:[Influence of chronic use of lead ions on rat spermatozoa]. 815 22

Epididymal sperm maturation culminates in the acquisition of functional competence by testicular spermatozoa. The expression of this functional state is dependent upon a redox-regulated, cAMP-mediated signal transduction cascade that controls the tyrosine phosphorylation status of the spermatozoa during capacitation. Analysis of superoxide anion (O2(-.)) generation by rat epididymal spermatozoa has revealed a two-component process involving electron leakage from the sperm mitochondria at complexes I and II and a plasma membrane NAD(P)H oxidoreductase. Following incubation in a glucose-, lactate-, and pyruvate-free medium (-GLP), O2(-.) generation was suppressed by 86% and 96% in caput and cauda spermatozoa, respectively. The addition of lactate, malate, or succinate to spermatozoa incubated in medium -GLP stimulated O2(-.) generation. This increase could be blocked by rotenone and oligomycin (R/O) in the presence of malate or lactate but not succinate. Stimulation with all three substrates, as well as spontaneous O2(-.) production in +GLP medium, was blocked by the flavoprotein inhibitor, diphenylene iodonium. Diphenylene iodonium, but not R/O, suppressed NAD(P)H-induced lucigenin-dependent chemiluminescence. This NAD(P)H-dependent enzyme resided in the sperm plasma membrane and its activity was regulated by zinc and uncharacterized cytosolic factors. Reverse transcription-polymerase chain reaction analysis indicated that the sperm NAD(P)H oxidoreductase complex is quite distinct from the equivalent leukocyte system.
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PMID:Analysis of reactive oxygen species generating systems in rat epididymal spermatozoa. 1156 31