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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Guanosine di- and triphosphates specifically decrease the affinity of chemotactic cAMP receptors in isolated Dictyostelium discoideum membranes. The K0.5 was increased from 50 nM to 150 nM. Receptors were shown to be heterogeneous in dissociation kinetics. In the absence of guanine nucleotides three dissociation processes could be resolved, having first order rate constants of 8.7 X 10(-4), 1.3 X 10(-2), and higher than 0.1 s-1. Guanine nucleotides decreased the affinity for cAMP by transforming the slowest dissociating receptor form (KD is 8 nM) to forms dissociating more rapidly. Our data indicate that a guanine nucleotide binding protein (G-protein) is involved in the transduction of the cAMP signal in D. discoideum.
Mol Cell Biochem 1985 Jul
PMID:Guanine nucleotides modulate the function of chemotactic cyclic AMP receptors in Dictyostelium discoideum. 299 88

Treatment of membranes from bovine cerebral cortex with N-ethylmaleimide (NEM) resulted in inhibition of gamma-aminobutyric acid (GABA) binding to GABAB receptors. The binding curve for increasing concentrations of agonist was shifted to the right by NEM treatment. Guanine nucleotide had little effect on the binding of GABA to NEM-treated membranes. The addition of purified GTP-binding proteins, which were the substrates of islet-activating protein (IAP), pertussis toxin, to the NEM-treated membranes caused a shift of the binding curve to the left, suggesting modification of GTP-binding proteins rather than receptors by NEM. Therefore, the effect of NEM on two purified GTP-binding proteins, Gi (composed of three subunits with molecular weight of alpha, 41,000; beta, 35,000; gamma, 10,000) and Go (alpha, 39,000; beta, 35,000; gamma, 10,000) was studied. NEM did not significantly change guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) binding and GTPase activity of these two proteins. In contrast, NEM-treated Gi and Go were not ADP-ribosylated by IAP and did not increase GABA binding to NEM-treated membranes. When alpha and beta gamma subunits were treated with NEM and then mixed with nontreated alpha and beta gamma to form Gi or Go, respectively, both oligomers with NEM-treated alpha-subunits lost their abilities to be IAP substrates and to couple to receptors. These results indicate that NEM uncoupled GTP-binding proteins from receptors by modifying alpha-subunits of GTP-binding proteins, and the site seemed to be on or near the site of ADP-ribosylation by IAP. When alpha and beta gamma subunits were treated with NEM and then mixed to form Gi or Go, GTP gamma S binding in the absence of Mg2+ and GTPase activity were changed, although they were not affected when oligomers were treated with NEM. The results suggest the existence of another sulfhydryl group which is protected from NEM by the association of subunits. The modification of this sulfhydryl group by NEM appeared to interfere with the interaction between alpha and beta gamma.
Mol Pharmacol 1986 Mar
PMID:Uncoupling of gamma-aminobutyric acid B receptors from GTP-binding proteins by N-ethylmaleimide: effect of N-ethylmaleimide on purified GTP-binding proteins. 300 32

We have studied the interaction of guanine nucleotides with alpha 1-adrenergic receptors of two cloned cell lines, the Madin Darby canine kidney (MDCK-D1) cells and BC3H-1 muscle cells. Although guanylylimidodiphosphate, Gpp(NH)p, had no effect on the affinity or the total number of [3H]prazosin-binding sites in membranes prepared from these cells, the nucleotide decreased the apparent affinity of the agonists (-)-epinephrine and (-)-norepinephrine in competing for [3H]prazosin-binding sites in both cell types. A maximal effect of Gpp(NH)p occurred at 10 microM. Guanine nucleotides were significantly more effective in shifting agonist affinity for the alpha 1 receptor than adenine nucleotides, and Mg2+ was required to observe a maximal effect. Binding of agonist to alpha 1-adrenergic receptors activated phosphatidylinositol (PI) hydrolysis in both cell types but had no effect on membrane adenylate cyclase activity. Incubation of MDCK cells for 19 hr with 100 ng/ml pertussis toxin, which eliminated the ability of pertussis toxin added to membranes to ADP-ribosylate 39-41-KDa substrate(s), failed to alter binding of agonists to alpha 1-adrenergic receptors, the ability of Gpp(NH)p to regulate agonist binding to these receptors, or epinephrine-stimulated PI hydrolysis and prostaglandin E2 production. Incubation of BC3H1 cells with pertussis toxin had no effect on the ability of epinephrine to stimulate PI turnover. These results show that binding of agonists to alpha 1-adrenergic receptors in mammalian kidney and muscle cells is regulated by guanine nucleotides, presumably by interaction with a guanine nucleotide-binding (G) protein. The failure of the G-protein to regulate adenylate cyclase activity and the lack of effect of pertussis toxin to alter receptor-mediated binding or functional activity suggests that a G-protein other than Gs, Gi, or Go interacts with alpha 1-adrenergic receptors in kidney and smooth muscle.
Mol Pharmacol 1987 Jan
PMID:Alpha 1-adrenergic receptor-linked guanine nucleotide-binding protein in muscle and kidney epithelial cells. 302 23

The characteristics of mu and delta opioid receptor sites present in human neuroblastoma SH-SY5Y cells were investigated using [D-Ala2-N-methyl-Phe4-Gly-(01)5]enkephalin (DAGO) and [2-D-penicillamine, 5-D-penicillamine]enkephalin (DPDPE), which are the most selective radioligands available for mu and delta sites, respectively. Scatchard analysis of the saturation isotherms revealed high affinity binding to a single class of sites for both [3H]DAGO (mu) and [3H]DPDPE (delta). [3H]DAGO labeled twice the number of sites compared to the binding capacity of [3H]DPDPE, yielding a mu/delta ratio of 2:1. Selective suppression of [3H]diprenorphine binding by specific opioid "blocking" ligands also showed a predominance of mu receptors, representing 65-70% of the total opioid sites. Competition binding studies carried out with a series of opiates and opioid peptides displayed higher potencies of mu- and delta-selective ligands in displacing the specific binding of [3H]DAGO and [3H]DPDPE, respectively. The [3H]diprenorphine/agonist competition curves were biphasic, indicating the high and low affinity states of mu and delta receptor sites in SH-SY5Y cells. Guanine nucleotide and sodium had differential effects on the agonist affinity and the proportion of high affinity states of mu and delta receptors. The mu and delta receptor sites were shown to be functionally coupled to adenylate cyclase. All of these data support the independent existence of mu and delta receptor types in human neuroblastoma cells. SH-SY5Y cells, therefore, represent a suitable model for investigating opioid-mediated responses in nerve cell populations.
Mol Pharmacol 1987 Jul
PMID:Comparative pharmacological properties and functional coupling of mu and delta opioid receptor sites in human neuroblastoma SH-SY5Y cells. 303 97

(R)-(-)-[77Br]4-Bromo-2,5-dimethoxyamphetamine [(R)-(-)-[77Br] DOB] was synthesized to a high specific activity (1875 +/- 50 Ci/mmol) and used to label membrane-associated recognition sites in rat brain. (R)-(-)-[77Br]DOB displayed high affinity (KD = 0.60 +/- 0.08 nM) for a relatively low density of binding sites (Bmax = 1.2 +/- 0.08 pmol/g of tissue) in rat cortical membranes as compared with [3H]ketanserin (KD = 0.65 +/- 0.1 nM; Bmax = 6.2 +/- 0.6 pmol/g of tissue). Guanine, but not adenine, nucleotides were found to inhibit specific (R)-(-)-[77Br]DOB binding. GTP (10(-4) M) did not eliminate specific (R)-(-)-[77Br]DOB binding but caused a competitive inhibition of the radioligand. Drug competition studies of 5-hydroxytryptamine (5-HT) and related agents indicate that both putative agonists and antagonists display nanomolar potency for these sites. A significant correlation (p less than 0.01) exists between drug potencies for (R)-(-)-[77Br]DOB-labeled sites and both 5-HT2 (r = 0.64) and 5-HT1C (r = 0.68) binding sites. However, the sites do not appear to be identical. Moreover, a significant correlation exists between drug potencies for (R)-(-)-[77Br]DOB-labeled sites and human hallucinogenic drug potencies (r = 0.89; p less than 0.01). We conclude that (R)-(-)-[77Br]DOB labels a unique 5-HT recognition site in rat brain that does not coincide with previously described 5-HT binding site subtypes. The (R)-(-)-[77Br]DOB site does not appear to be a high affinity "state" of the 5-HT2 receptor but may label a subset of heterogeneous 5-HT2 recognition sites.
Mol Pharmacol 1988 Oct
PMID:(R)-(-)-[77Br]4-bromo-2,5-dimethoxyamphetamine labels a novel 5-hydroxytryptamine binding site in brain membranes. 317 34

Different wild-type isolates of Dictyostelium discoideum exhibit extensive polymorphism in the length of restriction fragments carrying tRNA genes. These size differences were used to study the organisation of two tRNA gene families which encode a tRNA Val(GUU) and a tRNA Val(GUA) gene. The method used involved a combination of classical D. discoideum parasexual genetics and molecular genetics. The tRNA genes were mapped to specific linkage groups (chromosomes) by correlating the presence of polymorphic DNA bands that hybridized with the tRNA gene probes with the presence of genetic markers for those linkage groups. These analyses established that both of the tRNA gene families are dispersed among sites on several of the chromosomes. Information of nine tRNA Val(GUU) genes from the wild-type isolate NC4 was obtained: three map to linkage group I (C, E, F), two map to linkage group II (D, I), one maps to linkage group IV (G), one, which corresponds to the cloned gene, maps to either linkage group III or VI (B), and two map to one of linkage groups III, VI or VII (A, H). Six tRNA Val(GUA) genes from the NC4 isolate were mapped: one to linkage group I (D), two to linkage group III, VI or VII (B, C) and three to linkage group VII or III (A, E, F).
Mol Gen Genet 1987 Apr
PMID:Chromosomal mapping of tRNA genes from Dictyostelium discoideum. 347 95

A new high affinity antagonist photoaffinity crosslinking radioligand has been synthesized for use in studying adenosine receptors. This compound, PAPAXAC (8-[-4-[[[[[2-(4-aminophenylacetylamino)ethyl]amino]carbonyl]- methyl]oxy]phenyl]-1,3-dipropylxanthine), has been labeled with 125I by a chloramine T method. The radioligand [125I]PAPAXAC binds to A1 adenosine receptors from bovine cerebral cortex with high affinity (KD = 0.1 nM), appropriate stereoselectivity, and A1 adenosine receptor specificity. Binding is not perturbed by guanine nucleotides. Adenylate cyclase assays document that PAPAXAC is an antagonist capable of completely blocking the ability of N6-R-phenyl-2-propyladenosine (R-PIA) to inhibit adenylate cyclase activity via A1 adenosine receptors. [125I]PAPAXAC can be incorporated covalently into a peptide of Mr = 40,000 using the heterobifunctional crosslinking agent N-succinimidyl-6-(4'-azido-2'-nitrophenylamino)hexanoate. Covalent labeling can be inhibited with adenosine receptor ligands to demonstrate a potency series of R-PIA greater than S-PIA greater than NECA much greater than IBMX. Guanine nucleotides do not decrease covalent incorporation. These results suggest that antagonists such as [125I]PAPAXAC recognize the same A1 adenosine receptor-binding subunit as agonists, such as [125I]AZPNEA, which labels a similar Mr peptide with the same pharmacological potency series. This new antagonist photoaffinity crosslinking probe/radioligand should be of great utility in the molecular characterization of A1 adenosine receptors.
Mol Pharmacol 1987 Aug
PMID:A new high affinity, iodinated adenosine receptor antagonist as a radioligand/photoaffinity crosslinking probe. 361 92

Analysis of [3H]quinuclidinyl benzilate/acetylcholine competition curves indicated that the agonist acetylcholine bound with three different affinities to chick heart muscarinic receptors. The estimated KD values for acetylcholine were 2.7, 240, and 4000 nM. Mg2+ increased and guanosine 5'-(beta, gamma-imino)triphosphate (Gpp(NH)p) decreased the proportion of the receptors in the highest affinity state without altering the KD values. Monovalent cations increased the KD values of the three affinity states and obscured the detection of the highest affinity state. The nature of the three affinity states and the sites of action of Mg2+, guanine nucleotides, and monovalent cations were probed with three experimental protocols. Treatments with N-ethylmaleimide or pertussis toxin eliminated both the highest affinity state and the sensitivity to Gpp(NH)p. In contrast, partial effects of Mg2+ were retained after either of these treatments. The effects of monovalent cations on the affinity of the receptor for agonists were unaffected by both treatments. Solubilization of the receptors with digitonin-cholate yielded preparations displaying only the low affinity state for agonist. Agonist binding to the solubilized receptors was insensitive to Mg2+ and guanine nucleotides but retained sensitivity to monovalent cations. The results indicate that chick heart muscarinic receptors can exist in vitro in three agonist affinity states and that the entire population of receptors can be interconverted from one state to another by Mg2+ and guanine nucleotides. Guanine nucleotides presumably act via the inhibitory guanine nucleotide-binding regulatory (Ni) protein, whereas there appear to be at least two distinct sites of action of Mg2+. One site is associated with Ni. Another is distinguishable from Ni but does not appear to be on the receptor itself. The effect of monovalent cations on the interaction of agonists with cardiac muscarinic receptors is qualitatively different and mediated at distinct sites from the effects of Mg2+ and guanine nucleotides.
Mol Pharmacol 1985 Nov
PMID:Agonist interactions with cardiac muscarinic receptors. Effects of Mg2+, guanine nucleotides, and monovalent cations. 405 21

The binding of agonists and antagonists to Ri adenosine receptors of synaptosomal membranes from rat and bovine brain was studied. The effects of guanine nucleotides and temperature were analyzed with the aid of computerized curve fitting. Evidence is presented for two different states of the receptor: one of high and one of low affinity for agonists. Antagonists bind to both states with the same affinity. The two states are characterized by saturation, competition, and kinetic experiments with very similar results. Guanine nucleotides cause transition of the high- to the low-affinity state. The ratio of the KD values for the two affinity states is 90-150 in rat brain but only 10 in bovine brain. The proportions of the two affinity states are the same for all agonists tested; in the absence of exogenous guanine nucleotides, 75% of the total receptor population is in the high-affinity state, whereas in the presence of guanine nucleotides only 5% remain in the high-affinity state. Binding of antagonists to the receptor is enthalpy-driven whereas binding of the agonist (-)-N6-phenylisopropyladenosine to the high-affinity state of the receptor is entropy-driven. Binding of the agonist to the low-affinity state is enthalpy-driven and thus similar to the binding of antagonists. Our data indicate that guanine nucleotides convert the Ri adenosine receptor from a high- to a low-agonist affinity state and that agonist binding shows thermodynamic differences from antagonist binding only when it is to the high-affinity state of the receptor.
Mol Pharmacol 1984 Jul
PMID:Two affinity states of Ri adenosine receptors in brain membranes. Analysis of guanine nucleotide and temperature effects on radioligand binding. 608 14

The binding of [3H]quinuclidinyl benzilate ([3H]QNB) to cardiac muscarinic receptors was inhibited not only by classical muscarinic antagonists but also by nicotinic blocking agents and inhibitors of acetylcholinesterase. Gallamine, pancuronium, ambenonium, and decamethonium were the most potent of these agents examined. All of the nicotinic antagonists with significant muscarinic receptor activity had two or three quaternary nitrogens, and the potency of a series of these compounds was a function of the distance between quaternary nitrogens. The effects of gallamine and pancuronium were studied in detail because these neuromuscular blocking agents showed heterogeneity in their binding to cardiac muscarinic receptors, whereas classical muscarinic antagonists such as QNB and atropine did not. Gallamine did not compete for all of the [3H]QNB binding sites on atrial membranes, but left at least 20% of [3H]QNB binding unaffected. Curves of pancuronium competition for [3H]QNB binding were shallow, consistent with two binding sites for pancuronium, with approximately 20% having low affinity. Additionally, in the presence of gallamine or pancuronium, [3H]QNB binding sites were no longer homogeneous, and Scatchard plots became nonlinear. Guanine nucleotides did not alter the effect of gallamine or pancuronium on [3H]QNB binding. Gallamine and pancuronium showed no agonist activity but, like atropine, completely antagonized muscarinic receptor-mediated inhibition of cyclic AMP formation. However, differences in the behavior of gallamine and atropine suggested that gallamine was not a purely competitive antagonist. Gallamine did not protect against receptor alkylation by propylbenzilylcholine mustard, and [3H]QNB dissociation was apparently slowed by gallamine. We interpret our data to suggest that gallamine not only competes for [3H]QNB binding sites, but also binds at a secondary site on the receptor, forming a ternary complex with [3H]QNB. Heterogeneity in ligand binding is proposed to result from the dual actions of gallamine and pancuronium as ligands at both primary and secondary sites on the cardiac muscarinic receptor.
Mol Pharmacol 1983 Jul
PMID:Heterogeneity of binding sites on cardiac muscarinic receptors induced by the neuromuscular blocking agents gallamine and pancuronium. 613 50


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