We have compared the binding to the estrogen receptor (R) of different androgens and antiestrogens with their antiestrogenic activities on uterine growth. We found that estradiol (E2)1 and hydroxytamoxifen, a potent antiestrogen, displayed the same affinity for R. Conversely, androgens which have a much lower affinity for R and a much higher dissociation rate than E2, behave at high doses as full estrogens, with no significant antiestrogenic activity. We conclude that there is no correlation between the dissociation rate from R and the antiestrogenic activity of R ligands and that one cannot discriminate between estrogen and antiestrogen ligands by simply evaluating their binding to the cytosol R. 相似文献
Isolated adrenal cortex cells respond to the addition of ACTH1–39 or analogs with increased production of cyclic AMP and corticosterone. It is estimated that cyclic AMP production need proceed at less than 20% of maximum to induce maximum corticosterone production. ACTH1–24, [Lys17, Lys18]ACTH1–8 amide, and ACTH1–16 amide induce a maximum rate of cyclic AMP and of corticosterone production equal to those of ACTH1–39. The relative potencies as determined by cyclic AMP and by corticosterone production are in excellent agreement. The analog, ACTH5–24, induces maximum cyclic AMP production equal to 45% of that of the natural hormone, but as predicted, induces maximum corticosterone production equal to that of ACTH1–39. The derivative, [Trp(Nps)9]ACTH1–39 induces 77% of maximum corticosterone production and less than 1% of maximum cyclic AMP production. The fragment ACTH11–24 is a competitive antagonist of ACTH1–39 for both cyclic AMP and corticosterone production. The observations on agonists, a partial agonist and a competitive antagonist are in harmony with the “second messenger” role assigned to cyclic AMP. A provisional model, based on the fit of the experimental observations to a set of equations, provides expressions of “intrinsic activity,” “receptor reserve”, “sensitivity”, and “amplification” in terms of maximum cyclic AMP production, concentration of ACTH which induces maximum cyclic AMP production and concentration of cyclic AMP which induces maximum corticosterone production. 相似文献
Experiments were designed to determine whether or not various antiestrogens in direct competition with estradiol-17β (E2) would inhibit the translocation of the estrogen receptor complex from the cytoplasm to nuclei in rat uterine tissue. Incubation of the antiestrogens CI-628, -clomiphene, U-11,100A and MER-25 with rat uteri caused the nuclear uptake of the antiestrogen receptor complex which was greatest for most antiestrogens at concentrations of 1 × 10?6 to 1 × 10?5M. At higher concentrations of CI-628, -clomiphene, and U-11,100A the nuclear binding of the antiestrogen receptor complex was greatly decreased. Incubation of the antiestrogens with E2 resulted in a dramatic inhibition of the nuclear uptake of the estrogen receptor. -clomiphene, a weak estrogen, did not inhibit the movement of the uterine cytoplasmic receptor into the nuclear fraction. 相似文献
The reaction of almond β-glucosidase with -nitrophenyl-β--glucoside has been investigated over the temperature range +25° to ?45° using 50% aqueous dimethyl sulfoxide (DMSO) as solvent. At temperatures below those at which turnover occurs a “burst” of -nitrophenol proportional to the enzyme concentration is observed. Such a “burst” suggests the existence of a glucosyl-enzyme intermediate whose breakdown is rate-limiting, and provides a method for measuring the active-site normality. At pH 5.9, 25°, the presence of 50% DMSO causes an increase in Km from 1.7×10?3M (0%) to 1.7×10?2M, whereas Vmax is unchanged. The DMSO thus apparently acts as a competitive inhibitor with Ki = 0.7M. The Arrhenius plot for turnover is linear over the accessible temperature range with Ea = 23.0 ± 2.0 kcal/mole. 相似文献
The E0′ values for the conversion of horse heart cytochrome from the oxidized to the reduced form as a function of temperature have been measured in 0.10 M NaCl, 0.10 M sodium phosphate, pH 7.0 solutions in H2O and D2O. In H2O, the decrease in the E0′ value is linear with increasing temperature up to 42°C. Above this temperature, the decrease is again linear but with a much greater slope. In D2O solutions, however, this biphasic behavior was not observed but instead a single line was obtained over the temperature range studied (25°C to 50°C). These results are interpreted in terms of the ability of NaCl to cause a destructuring of the bulk H2O above 42°C but not in the more stable D2O (Kreishman, Foss, Inoue and Leifer (1976) , 5431–5435). This decrease in water structure results in a shift in the equilibrium to the larger oxidized form as indicated by the decrease in E0′. 相似文献
Previous proton nuclear magnetic resonance (nmr) studies have indicated that inositol hexaphosphate (IHP) can stabilize hemoglobin (Hb) Kansas in a deoxy-like quaternary structure even when fully liganded with carbon monoxide (CO) (S. Ogawa, A. Mayer, and R. G. Shulman, 1972, Biochem. Biophys. Res. Commun., 49, 1485–1491). In the present report we have investigated both CO binding at equilibrium and the CO binding and release kinetics to determine if Hb Kansas + IHP is devoid of cooperativity, as would be suggested by the nmr studies just quoted. The equilibrium measurements show that Hb Kansas + IHP has a very low affinity for CO ( Hg and Keq = 5.4 × 105M?1) and almost no cooperativity (n = 1.1) at pH 7, 25 °C. The CO “on” and “off” kinetics also show no evidence for cooperativity. In addition, the equilibrium constant estimated from the kinetic rate constants (Keq = 5.2 × 105M?1 with kon = 1.03 × 105M?1 · S? and koff = 0.198 S?1) is in excellent agreement with the equilibrium constant determined directly. Thus, both kinetic and equilibrium measurements allow us to conclude that CO binding to Hb Kansas + IHP occurs without significant cooperativity. 相似文献
Two enzymes involved in the biosynthesis of riboflavin have been found in , an organism that produces and excretes riboflavin in large quantities. The first of these (called “reductase”) functions to reduce the ribose group of 2,5-diamino-6-oxy-4-(5′-phosphoriboyslamino)pyrimidine (PRP, Compound II, Fig. 1) to the ribityl group of Compound IV (Fig. 1). The second enzyme (called “deaminase”) catalyzes the deamination of Compound IV to Compound V. The evidence indicates that in the reductase functions before the deaminase, in contrast to the reaction sequence known to operate in in which deamination takes place before reduction (Burrows, R.B. and Brown, G.M. (1978) J. Bacteriol., , 657–667). 相似文献
The steroid hormone-receptor complex concentrations measured by “differential dissociation” techniques have to be corrected to obtain the true concentrations of receptor binding sites (Bs). For the calculation of Bs, the parameters kn (product of the equilibrium association constant and the concentration of binding sites of the “nonspecific” component) and f (fraction of the nonspecific binding measured in the experimental estimates of bound ligand by a given technique), previously proposed by Blondeau and Robel (J. P. Blondeau and P. Robel, 1975, Eur. J. Biochem.55, 375–384) are important. A new parameter of interest, ? [], is discussed. The measurement of this parameter ? for three “differential dissociation” techniques allows the comparison of their efficiency and their reliability under various conditions for hormone receptor measurement in cytosol. Charcoal and hydroxylapatite methods are more efficient than the Sephadex G-25 filtration method. It is demonstrated that the “isotopic dilution” correction generally used for the estimation of the background of a given technique may be incorrect whatever the method of correction. A new method, the “double concentration measurement,” is developed. This method is simple, rapid, and precise. It requires two receptor binding measurements at two different saturating concentrations of ligand. This method allows the measurement of the estradiol receptor binding activity from calf uterine cytosol, with an error of less than 5% in samples containing the receptor either free or previously complexed with radioactive hormone, even in the presence of very high concentrations (≤0.5 μm) of radioactive steroid. 相似文献
Adenosine has a significant role in many functions of the central nervous system. Behaviorally, adenosine and adenosine analogs have marked depressant effects. Electrophysiologically, adenosine reduces spontaneous neuronal activity and inhibits transsynaptic potentials interaction with extracellular receptors. Biochemically, adenosine inhibits adenylate cyclase a “high” affinity receptor, and activates adenylate cyclase a “low” affinity receptor. These receptors, called “A1” and “A2” respectively, show differing profiles for activation by adenosine analogs. Radioactive N6-cyclohexyladenosine binds selectively to the “high” affinity receptor. One major class of antagonists is known at adenosine receptors: the alkylxanthines, including caffeine and theophylline. Radioactive 1,3-diethyl-8-phenylxanthine, a particularly potent antagonist, appears to bind to both low and high affinity adenosine receptors. Behavioral, electrophysiological, and biochemical effects of alkylxanthines are consistent with the hypothesis that the central stimulatory actions of caffeine and theophylline are due in large part to antagonism of central adenosine receptors. 相似文献
The synthesis of -4,5,6-trinor-3,7-inter--phenylene-3-oxaprostaglandins oxaprostaglandins of the E1 and F1α series7 from 6-endo-(1-heptenyl)-bicyclo[3:1:0]hexan-3-one (III), is described. Preliminary biological screening data for gerbil colon smooth muscle stimulation, rat blood pressure and substrate specificity toward 15-hydroxyprostaglandin dehydrogenase is presented. Platelet function studies, both and of -4,5,6-trinor-3,7-inter--phenylene-3-oxa-PGE1, methyl ester (VIII) are presented. 相似文献
ATPase preparations from rat brain, dog kidney, and human red blood cells also catalyze a K+-dependent phosphatase reaction. K+ activation and Na+ inhibition of this reaction are described quantitatively by a model featuring isomerization between E1 and E2 enzyme conformations with activity proportional to E2K concentration: Differences between the three preparations in for K+ activation can then be accounted for by differences in equilibria between E1K and E2K with dissociation constants identical. Similarly, reductions in produced by dimethyl sulfoxide are attributable to shifts in equilibria toward E2 conformations. Na+ stimulation of K+-dependent phosphatase activity of brain and red blood cell preparations, demonstrable with KCl under 1 mM, can be accounted for by including a supplementary pathway proportional to E1Na but dependent also on K+ activation through high-affinity sites. With inside-out red blood cell vesicles, K+ activation in the absence of Na+ is mediated through sites oriented toward the cytoplasm, while in the presence of Na+ high-affinity K+-sites are oriented extracellularly, as are those of the ATPase reaction. Dimethyl sulfoxide accentuated Na+-stimulated K+-dependent phosphatase activity in all three preparations, attributable to shifts from the E1P to E2P conformation, with the latter bearing the high-affinity, extracellularly oriented K+-sites of the Na+-stimulated pathway. 相似文献
The presence of two high affinity estrogen binding proteins in the uterine cytosol of the immature rat has been observed.Besides the 8 S cytosol estrogen , there is a 4–5 S fraction binding estradiol and estrone with a large capacity. In fact, the two binding systems have a different affinity for estradiol and estrone, the receptor binding more the former and the 4–5 S fraction more the latter. Exposure of the cytosol to specific anti-α1-Fetoprotein antibodies suppresses a large part of the 4–5 S binder, if not the totality. Moreover the estrogen binding 4–5 S fraction decreases with increasing age until puberty, while the increases. These results suggest therefore that the estradiol-estrone binding 4–5 S peak of the uterine cytosol is mainly made up of Estradiol Binding Plasma Protein-α1-Fetoprotein (EBP-AFP). Also they confirm that “cytosol” should be taken as an operational fraction which may include extracellular components.During the course of these experiments, it has been observed that the increase of the estradiol is more rapid than the other uterine cytosol proteins until the 8th day, and that there is a second period of growth when it follows the development of the uterus and of the animal, as if it had reached a constant number of binding sites per cell. 相似文献
Various hemoglobin derivatives have been labeled at the Cys-β93 residue with a bulky and “strongly immobilized” nitroxide maleimide (I) and a smaller, more flexible and “weakly immobilized” nitroxide iodoacetamide (II) and crystallized. The angular dependence of the paramagnetic resonance of the spin-label was measured for the ab, and planes at 298 K and 77 K for spin-labeled crystals of Oxyhemoglobin, methemoglobin fluoride, and methemoglobin azide. In the case of the methemoglobin crystals, the angular variation of the heme resonance was also monitored at 77 K. From the hyperfine splitting data, the spin-label I was found to assume specific orientations at both temperatures with some motional narrowing at 298 K. Spin-label II is specifically oriented only at room temperature but is frozen at 77 K in random orientations. Oxyhemoglobin labeled with I (I-HbO22) has the most prominent spin-label orientation (z∥b, x∥a) and the less abundant spin-labels with (z∥b ± 15 °) (Ohnishi et al., 1966). The corresponding spin-label orientations for I-Hb+ F? are () and (). Crystals of I-Hb+ N?3 have spin-labels oriented along angular directions similar, but not identical to those of I-Hb+F?. Therefore, there are probably significant peptide segmental displacements when HbO2 is oxidized to methemoglobins. At 25 °C II-Hb+ N?3 has spin-label orientations not too different from those in I-Hb+ N?3, whereas in HbO2 the two spin-labels show significant differences in their orientations. 相似文献