Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have characterized the ANF-R2 receptor-mediated inhibition of adenylate cyclase with respect to its modulation by several regulators. ANF (99-126) inhibits adenylate cyclase activity only in the presence of guanine nucleotides. The maximal inhibition (approximately 45%) was observed in the presence of 10-30 microM GTP gamma S, and at higher concentrations, the inhibitory effect of ANF was completely abolished. ANF-mediated inhibition was not dependent on the presence of monovalent cations, however Na+ enhanced the degree of inhibition by about 60%, whereas K+ and Li+ suppressed the extent of inhibition by about 50%. On the other hand, divalent cation, such as Mn2+ decreased the degree of inhibition in a concentration dependent manner, with an apparent Ki of about 0.7 mM, and at 2 mM; the inhibition was completely abolished. In addition, proteolytic digestion of the membranes with trypsin (40 ng/ml) resulted in the attenuation of ANF-mediated inhibition of adenylate cyclase. Other membrane disrupting agents such as neuraminidase and phospholipase A2 treatments also inhibited completely, the ANF-mediated inhibition of enzyme activity. N-Ethylmaleimide (NEM), phorbol ester and Ca(2+)-phospholipid dependent protein kinase (C-kinase) which have been shown to interact with inhibitory guanine nucleotide regulating protein (Gi) also resulted in the attenuation of ANF-mediated inhibition of adenylate cyclase activity. These results indicate that in addition to the Gi, the phospholipids and glycoproteins may also play an important role in the expression of ANF-R2 receptor-mediated inhibition of adenylate cyclase.
Mol Cell Biochem 1992 Jul 06
PMID:Characterization of ANF-R2 receptor-mediated inhibition of adenylate cyclase. 132 94

We have shown previously that cultured human lung cancer cells of different histologic types express multiple opioid receptors that can regulate their growth. In this report, we show that these cells also express specific, saturable, and high-affinity binding sites (Kd approximately 1 nM) for the non-opioid phencyclidine (PCP), [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,b]cyclohepten-5,10-imine hydrogen maleate] (MK-801) and sigma N-allylnormetazocine (SKF-10,047) receptor ligands. Characterization of these binding sites showed them to be protein in nature and sensitive to the guanine nucleotide GTP. Pharmacological studies showed that (+) MK-801 and (+) SKF-10,047 competed with each other for their binding sites and also for the methadone binding site present in these cells. However, the mu and delta opioid ligands did not compete for (+) MK-801 and (+) SKF-10,047 binding sites. In addition, these binding sites on lung cancer cells appear to be distinct from the N-methyl D-aspartate/PCP receptor ionophore complex reported to be present in rat brain. MK-801 and SKF-10,047, at nM concentrations, were found to inhibit the growth of these cells in culture within a few hours of exposure, and this effect was irreversible after 24 h. The growth effects of these ligands could not be reversed by the opioid antagonist naloxone, suggesting involvement of nonopioid type receptors in the actions of these ligands. The abundant expression of biologically active MK-801 and SKF-10 047 binding sites in these cell lines, distinct from those in rat brain, suggests that these cell lines may prove to be a valuable source for further characterization and purification of these binding sites.
Mol Biol Cell 1992 Jun
PMID:Biologically active MK-801 and SKF-10,047 binding sites distinct from those in rat brain are expressed on human lung cancer cells. 132 49

Previous studies have indicated that ADP-glucose pyrophosphorylase (ADPGlc PPase) from the cyanobacterium Anabaena sp. strain PCC 7120 is more similar to higher-plant than to enteric bacterial enzymes in antigenicity and allosteric properties. In this paper, we report the isolation of the Anabaena ADPGlc PPase gene and its expression in Escherichia coli. The gene we isolated from a genomic library utilizes GTG as the start codon and codes for a protein of 48,347 Da which is in agreement with the molecular mass determined by SDS-PAGE for the Anabaena enzyme. The deduced amino acid sequence is 63, 54, and 33% identical to the rice endosperm small subunit, maize endosperm large subunit, and the E. coli sequences, respectively. Southern analysis indicated that there is only one copy of this gene in the Anabaena genome. The cloned gene encodes an active ADPGlc PPase when expressed in an E. coli mutant strain AC70R1-504 which lacks endogenous activity of the enzyme. The recombinant enzyme is activated and inhibited primarily by 3-phosphoglycerate and Pi, respectively, as is the native Anabaena ADPGlc PPase. Immunological and other biochemical studies further confirmed the recombinant enzyme to be the Anabaena enzyme.
Plant Mol Biol 1992 Oct
PMID:Molecular cloning and expression of the gene encoding ADP-glucose pyrophosphorylase from the cyanobacterium Anabaena sp. strain PCC 7120. 132 5

We previously suggested that gonadotropin-releasing-hormone (GnRH) analogues activate the phosphoinositide pathway in rat mammary tumor membranes. In the present study we analyzed the binding of GnRH analogues to these membranes and assessed its modulation by guanine nucleotides. [125I]Buserelin (a GnRH superagonist) binding is specific because it is displaced only by GnRH analogues. Scatchard plot analysis reveals high affinity binding sites (Kd = 2.5 +/- 0.8 nM, Bmax = 250 +/- 120 fmol/mg membrane protein) and low affinity binding sites (Kd 1.1 +/- 0.3 microM, Bmax = 200 +/- 105 pmol/mg membrane protein). Guanine nucleotides increased the ED50 of [125I]buserelin displacement, and almost completely eliminated the high affinity binding. Similar results were obtained with [125I]D-Trp6-GnRH--another GnRH superagonist. The inhibition of buserelin binding by guanine nucleotides was specific for nucleotides that interact with G-binding proteins and was dose-dependent with a maximal effect at 10 microM GTP gamma S. Kinetic analysis of buserelin binding revealed that the dissociation rate increased at least 4-fold in the presence of 10 microM GTP gamma S. These results support the hypothesis that GnRH analogues interact directly with mammary tumors and activate a G-protein-dependent transducing mechanism.
Mol Cell Endocrinol 1992 May
PMID:Guanine nucleotide modulation of high affinity gonadotropin-releasing hormone receptors in rat mammary tumors. 132 49

To gain insight into the molecular mechanism underlying melatonin binding and signal transduction in the chick brain, we have investigated the role of -SH groups, using a sulfhydryl alkylating reagent N-ethylmaleimide (NEM). At least two -SH groups are involved in the formation of the receptor-G protein complex: one is sensitive to and the other relatively insensitive to NEM. Alkylation of the sensitive group selectively abolishes high affinity binding of 2-[125I]iodomelatonin ([125I]MEL), similar to the effect induced by GTP, thus leading to a complete loss of sensitivity to nucleotides. Modification of both groups causes a marked reduction in binding capacity. Agonists with high affinity, but not other compounds with low affinity for the melatonin receptor, protect against alkylation by NEM. GTP gamma s does not significantly alter the reactivity of -SH groups towards NEM, but agonist-protected receptors remain sensitive to this nucleotide. Moreover, NEM pretreatment blocks the inhibitory effect of melatonin on forskolin-stimulated adenylate cyclase activity in chick brain. These data suggest that the -SH group modulating agonist affinity may lie within the coupling domain between the receptor and G protein but outside of the GTP binding site. In addition, sulfhydryl groups are essential for melatonin binding and signal transduction in chick brain.
Mol Cell Endocrinol 1992 May
PMID:Involvement of multiple sulfhydryl groups in melatonin signal transduction in chick brain. 132 52

In mammalian cells, the signal recognition particle (SRP) receptor is required for the targeting of nascent secretory proteins to the endoplasmic reticulum (ER) membrane. We have identified the Saccharomyces cerevisiae homologue of the alpha-subunit of the SRP receptor (SR alpha) and characterized its function in vivo. S. cerevisiae SR alpha is a 69-kDa peripheral membrane protein that is 32% identical (54% chemically similar) to its mammalian homologue and, like mammalian SR alpha, is predicted to contain a GTP binding domain. Yeast cells that contain the SR alpha gene (SRP101) under control of the GAL1 promoter show impaired translocation of soluble and membrane proteins across the ER membrane after depletion of SR alpha. The degree of the translocation defect varies for different proteins. The defects are similar to those observed in SRP deficient cells. Disruption of the SRP101 gene results in an approximately sixfold reduction in the growth rate of the cells. Disruption of the gene encoding SRP RNA (SCR1) or both SCR1 and SRP101 resulted in an indistinguishable growth phenotype, indicating that SRP receptor and SRP function in the same pathway. Taken together, these results suggest that the components and the mechanism of the SRP-dependent protein targeting pathway are evolutionarily conserved yet not essential for cell growth. Surprisingly, cells that are grown for a prolonged time in the absence of SRP or SRP receptor no longer show pronounced protein translocation defects. This adaptation is a physiological process and is not due to the accumulation of a suppressor mutation. The degree of this adaptation is strain dependent.
Mol Biol Cell 1992 Aug
PMID:Signal recognition particle receptor is important for cell growth and protein secretion in Saccharomyces cerevisiae. 132 99

To probe for the involvement of Ca2+/calmodulin-dependent protein kinase II in the regulation of insulin secretion, the effects of a specific inhibitor of this enzyme, KN-62, on secretagogue-stimulated insulin secretion, cytosolic Ca2+ concentration ([Ca2+]i) rise, membrane depolarization, and nutrient metabolism were examined in HIT-T15 cells. KN-62 dose-dependently inhibited insulin secretion induced by a nutrient mixture (10 mM glucose, 5 mM leucine, and 5 mM glutamine) alone or combined with either the Ca(2+)-mobilizing receptor agonist bombesin or the cAMP-raising agent forskolin in intact cells. KN-62 did not affect Ca(2+)- or GTP analogue-induced insulin secretion from permeabilized cells, indicating an action at a step before exocytosis. The stimulating effects of nutrients on insulin secretion, [Ca2+]i, and membrane depolarization were potentiated by bombesin. Similarly, bombesin promoted a larger depolarization and [Ca2+]i rise in the presence of nutrients. This was associated with enhanced Ca2+ mobilization and the appearance of sustained [Ca2+]i elevation. The bombesin-induced membrane depolarization, like the nutrient effect, was inhibited by diazoxide, suggesting that this is due to closure of ATP-sensitive K+ channels. Bombesin elicited Ca2+ influx by both membrane potential-sensitive and -insensitive conductance pathways. KN-62 did not affect Ca2+ mobilization and only partially reduced Ca2+ entry during the sustained [Ca2+]i rise in bombesin-stimulated cells. When added before or during the stimulation, KN-62 dose-dependently inhibited nutrient- and KCl-stimulated [Ca2+]i elevation and Mn2+ influx (reflecting Ca2+ entry). The calmodulin antagonist CGS 9343B and the L-type Ca2+ channel blocker SR-7037 mimicked the inhibitory effect of KN-62 on stimulated insulin secretion and [Ca2+]i elevation. Membrane depolarization and nutrient metabolism (reduction of a tetrazolium derivative), however, were not altered by KN-62 treatment, indicating that the early coupling events from nutrient metabolism to closure of ATP-sensitive K+ channels remain operative. These results suggest that KN-62 and the calmodulin antagonist CGS 9343B inhibit Ca2+ influx by means of direct interaction with L-type Ca2+ channels, which, in turn, causes inhibition of stimulated insulin secretion. Thus, it appears that Ca2+/calmodulin-dependent protein kinase II is not involved in the regulation of insulin secretion.
Mol Pharmacol 1992 Sep
PMID:Inhibition of voltage-gated Ca2+ channels and insulin secretion in HIT cells by the Ca2+/calmodulin-dependent protein kinase II inhibitor KN-62: comparison with antagonists of calmodulin and L-type Ca2+ channels. 132 47

The discovery of mutated, GTPase-deficient alpha subunits of Gs or Gi2 in certain human endocrine tumors has suggested that heterotrimeric G proteins play a role in the oncogenic process. Expression of these altered forms of G alpha s or G alpha i2 proteins in rodent fibroblasts activates or inhibits endogenous adenylyl cyclase, respectively, and causes certain alterations in cell growth. However, it is not clear whether growth abnormalities result from altered cyclic AMP synthesis. In the present study, we asked whether a recently discovered family of G proteins, Gq, which does not affect adenylyl cyclase activity, but instead mediates the activation of phosphatidylinositol-specific phospholipase C harbors transforming potential. We mutated the cDNA for the alpha subunit of murine Gq in codons corresponding to a region involved in binding and hydrolysis of GTP. Similar mutations unmask the transforming potential of p21ras or activate the alpha subunits of Gs or Gi2. Our results show that when expressed in NIH 3T3 cells, activating mutations convert G alpha q into a dominant acting oncogene.
Mol Cell Biol 1992 Oct
PMID:Mutated alpha subunit of the Gq protein induces malignant transformation in NIH 3T3 cells. 132 59

Agonist binding to guanine nucleotide-binding protein (G protein)-coupled receptors in membranes of myeloid differentiated human leukemia (HL-60) cells is inhibited by guanine nucleotides, most potently by the GTP analog guanosine 5'-(gamma-thio)triphosphate (GTP gamma S). In order to study whether GTP gamma S formed locally from adenosine 5'-(gamma-thio)triphosphate (ATP gamma S) and GDP by nucleoside diphosphokinase has any advantage over exogenously added GTP gamma S in binding to and activating G proteins, regulation of complement component 5a (C5a) binding to its receptors, as well as formation of GTP gamma S, was studied in membranes of HL-60 cells. GTP gamma S added to HL-60 membranes potently inhibited binding of 125I-C5a (IC50 about 3 nM), an effect not influenced by addition of either GDP or ATP gamma S. When HL-60 membranes were incubated with the combination of ATP gamma S and GDP, a marked potentiation (up to 300-fold) of the inhibition caused by either GDP or ATP gamma S alone was observed. By measuring nucleoside diphosphokinase-catalyzed formation of GTP gamma S and inhibition of 125I-C5a binding in the presence of GDP and ATP gamma S under identical assay conditions, it was found that formed GTP gamma S inhibited binding of 125I-C5a with an IC50 value of about 0.3 nM, thus being about 10-fold more potent than exogenously added GTP gamma S. These data suggest that the GTP gamma S-forming nucleoside diphosphokinase is closely associated with the C5a receptor-G protein complex and channels the formed GTP gamma S into the G protein.
Mol Pharmacol 1992 Nov
PMID:Evidence for nucleoside diphosphokinase-dependent channeling of guanosine 5'-(gamma-thio)triphosphate to guanine nucleotide-binding proteins. 133 59

Bovine pulmonary artery smooth muscle (SM) cells express a novel 5-hydroxytryptamine (5-HT) (5-HT4-like) receptor coupled to cAMP accumulation. cAMP radioimmunoassay established the agonist and antagonist profiles of this receptor. 5-HT (EC50 = 91 +/- 33 nM) and 5-methoxytryptamine were equipotent at the SM cell 5-HT receptor and both were more potent than 5-carboxamidotryptamine. Other tryptamine derivatives were less potent but remained full agonists. These findings are consistent with previous reports regarding 5-HT4 and 5-HT4-like receptors in the central nervous system. The most potent antagonists were the antidepressant compounds nortriptyline (IC50 = 177 +/- 153 nM) and zimelidine (IC50 = 202 +/- 101 nM). The 5-HT3 and 5-HT4 antagonist 3-tropanyl-indole-3-carboxylate (ICS 205-930) was also a competitive antagonist at this 5-HT4-like receptor (pA2 = 6.3). Antagonist affinities differed slightly at the SM cell receptor, compared with other 5-HT4 and 5-HT4-like receptors in the central nervous system. Nonetheless, the SM cell 5-HT4-like receptor displayed the same differential antagonist potencies as reported for these other receptors (ICS 205-930 > MDL 72222 and mianserin > ketanserin). 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) was the most potent agonist for this 5-HT4-like receptor (EC50 = 6.4 +/- 3.4 nM). 8-OH-DPAT-induced cAMP accumulation could be blocked by ICS 205-930 but not by the 5-HT1A antagonist 1-(2-methoxyphenyl)-4-[4-(2-pthalimido)butyl]piperazine hydrobromide, distinguishing the SM cell 5-HT receptor from 5-HT1A receptors. The mechanism of 5-HT-stimulated cAMP production was also investigated. First, GTP augmented basal and 5-HT-stimulated cAMP accumulation. Second, antisera to the carboxyl terminus of the alpha subunit of Gs, attenuated 5-HT-mediated adenylate cyclase activation. This established that 5-HT-stimulated cAMP accumulation in SM cells required GS. These findings suggest that SM cells express a novel 5-HT4-like receptor positively coupled to adenylate cyclase. An unexpected finding was that 8-OH-DPAT is a potent partial agonist. These studies suggest that there may be heterogeneity among 5-HT4-like receptors.
Mol Pharmacol 1992 Nov
PMID:8-hydroxy-2-(di-n-propylamino)tetralin-responsive 5-hydroxytryptamine4-like receptor expressed in bovine pulmonary artery smooth muscle cells. 133 64


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