Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: CAS:7665-99-8 (cGMP)
 21,074 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pure cGMP-inhibited cAMP phosphodiesterase (cGI-PDE) in micrograms quantities was isolated from bovine aortic smooth muscle after more than 5000-fold purification using DEAE ion-exchange and affinity chromatography with a derivative of the specific cGI-PDE inhibitor cilostamide conjugated as a ligand to aminoethyl agarose (CIT-agarose). The cGI-PDE, which constituted about half of the high affinity cAMP-PDE activity of a tissue homogenate, was identified with a 105-kDa protein on SDS-PAGE through use of antibodies towards the human platelet, bovine cardiac and bovine adipose tissue cGI-PDE in Western blot and immunoprecipitation/immunoinactivation analysis. As observed during purification of the enzyme from other tissues the enzyme protein was exquisitely sensitive to proteolytic nicking during purification, resulting in several 30-77-kDa polypeptide fragments. Rapid immunoprecipitation from fresh tissue extracts was the only was found to partially prevent the proteolysis. The native enzyme had apparent molecular sizes of approx. 100,000 or, mainly approx. 220,000 by gel chromatography, presumably indicating the presence of monomeric and dimeric forms. The enzyme hydrolyzed cAMP and cGMP with normal Michaelis-Menten kinetics with Km of 0.16 and 0.09 microM, respectively, with Vmax for hydrolysis of cAMP of 0.3 compared to 3.1 mumol/min per mg protein for cAMP. The enzyme was potently and selectively inhibited by cGMP (IC50 approximately 0.25 microM) and the cardiotonic/vasodilatory drugs OPC-3911 (a cilostamide derivative), milrinone and CI-930 (IC50 approximately 0.05, 0.40 and 0.25 microM, respectively). The cGI-PDE was phosphorylated by cAMP-dependent protein kinase as has been reported for the analogous enzymes in heart, adipose tissue and platelets. The identification of a cGI-PDE in the aortic smooth muscle and its inhibitor specificity is consistent with the hypothesis that inhibition of this enzyme is important in the mechanism through which these drugs produce vasorelaxation.
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PMID:Purification and properties of the cGMP-inhibited cAMP phosphodiesterase from bovine aortic smooth muscle. 131 3

We have cloned a cDNA for a myocardial cGMP-inhibited cAMP phosphodiesterase (cGI PDE) from a human heart cDNA library in lambda Zap II. The open reading frame [3.5 kilobases (kb)] of cDNA clone n.13.2 (7.7 kb) encodes a protein of 125 kDa. In Northern blots of total human ventricle RNA, a single mRNA species (8.3 kb) hybridized with a 4-kb EcoRI restriction fragment of clone n.13.2 cDNA (containing the entire open reading frame). The carboxyl-terminal region of the deduced amino acid sequence of the cGI PDE contains the putative catalytic domain conserved among mammalian PDE families. A partial cDNA clone, n.2, encoding a truncated, 54-kDa cGI PDE containing the conserved domain was expressed as a catalytically active fusion protein in Escherichia coli. cAMP hydrolytic activity was inhibited by cGMP and OPC 3911 but not by rolipram. Thus, this report provides direct proof that the conserved domain contains the catalytic core of cGI PDEs.
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PMID:Molecular cloning and expression of human myocardial cGMP-inhibited cAMP phosphodiesterase. 131 35

The heterotrimeric guanine nucleotide binding proteins (G proteins) are activated by sensory or hormone receptors. In turn, the G proteins activate effector proteins such as adenylyl cyclase, cyclic guanosine 3',5'-monophosphate phosphodiesterase (cGMP PDE), phospholipase C, and potassium and calcium ion channels by mechanisms that are poorly understood. A site on the alpha subunit of the G protein transducin (alpha t) has been identified that interacts with and activates cGMP phosphodiesterase, the effector enzyme in rod photoreceptors. A 22-amino acid peptide, corresponding to residues 293 to 314 from the COOH-terminal region of alpha t, fully mimicked alpha t and potently activated PDE. This region is adjacent to the receptor activation domain; thus, the alpha subunit of this G protein has a site for interaction with both its effector and receptor that maps near the COOH-terminus.
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PMID:A site on rod G protein alpha subunit that mediates effector activation. 131 58

Insulin induced phosphorylation and activation of the cGMP inhibited cAMP phosphodiesterase (cGI-PDE) in human platelets were demonstrated after isolation of the enzyme with specific polyclonal cGI-PDE antibodies. The demonstration of this insulin effect required suppression of basal cGI-PDE phosphorylation, through the use of the protein kinase inhibitor H-7 (1-(5-isoquinolinylsulfonyl)-2-methylpiperazine). The human platelet insulin receptor beta-subunit, previously identified as a 97 kDa polypeptide, was detected with the use of wheat germ agglutinin chromatography and anti-phosphotyrosine antibodies. These results suggest that insulin, through phosphorylation/activation of cGI-PDE, could decrease cAMP/cAMP dependent protein kinase (cAMP-PK) activity and thereby make the platelets more sensitive towards aggregating agents.
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PMID:Insulin induced phosphorylation and activation of the cGMP-inhibited cAMP phosphodiesterase in human platelets. 132 13

8-(p-Chlorophenylthio)-cGMP (8-pCPT-cGMP) and 8-bromo-cGMP were compared with respect to their chemical and biological properties in order to evaluate their potential as selective activators of cGMP-dependent protein kinase (cGMP-PK; EC 2.7.1.37) in intact human platelets. 8-pCPT-cGMP, 8-Br-cGMP and cGMP were shown to be potent and selective activators of purified bovine lung cGMP-PK and of cGMP-PK present in human platelet membranes when compared with the activation of cAMP-dependent protein kinase (cAMP-PK; EC 2.7.1.37). 8-pCPT-cGMP was not hydrolysed by the purified cGMP-stimulated phosphodiesterase (cGS-PDE), cGMP-inhibited phosphodiesterase (cGI-PDE) and Ca(2+)-calmodulin-dependent phosphodiesterase (CaM-PDE), whereas cGMP and, to a lesser extent, 8-Br-cGMP were hydrolysed by all three types of 3',5' cyclic nucleotide phosphodiesterases (EC 3.1.4.17) examined. Also, 8-pCPT-cGMP was not hydrolysed by a human platelet homogenate which contains a high level of the cGMP-specific cGMP-binding phosphodiesterase (cGB-PDE). Additionally, 8-pCPT-cGMP did not activate the cGS-PDE or inhibit the cGI-PDE, whereas half-maximal inhibition of cGI-PDE occurred at 8 microM 8-Br-cGMP. The apparent lipophilicity of 8-pCPT-cGMP was higher than that of 8-Br-cGMP. Extracellular application of 8-pCPT-cGMP to intact human platelets reproduced the pattern of protein phosphorylation induced by sodium nitroprusside (SNP), a cGMP-elevating inhibitor of platelet activation. Quantitatively, 8-pCPT-cGMP was more effective than 8-Br-cGMP in inducing phosphorylation of the 46/50 kDa vasodilator-stimulated phosphoprotein, a major substrate of cGMP-PK in intact platelets. As observed with SNP, pretreatment of human platelets with 8-pCPT-cGMP prevented the aggregation induced by thrombin. The results suggest that 8-pCPT-cGMP is a very potent and selective activator of cGMP-PK in cell extracts and in intact human platelets and, in this respect, is superior to 8-Br-cGMP and other cGMP analogs used for intact cell studies. The data also suggest that inhibition of platelet activation in intact human platelets by nitrovasodilators is mediated by cGMP-PK.
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PMID:Analysis of the functional role of cGMP-dependent protein kinase in intact human platelets using a specific activator 8-para-chlorophenylthio-cGMP. 132 24

We have identified and sequenced cDNA clones that encode for the human beta-subunit of rod cGMP phosphodiesterase (PDEB). A single 2565-bp open reading frame that codes for an 854-amino-acid protein was identified. The human beta-subunit protein is 90% identical to the bovine beta-subunit and 91% identical to the mouse protein. Northern blot analysis indicates that the gene is expressed as an abundant 3.5-kb transcript in retina and as a rare 2.9-kb transcript in brain. The isolation of cDNAs from human brain cDNA libraries confirms the brain as a site of expression for this gene. The molecular defect underlying retinal degeneration in the rd mouse has been found to be a nonsense mutation in the beta-subunit of the mouse cGMP PDE, resulting in a truncated protein (Pittler et al., 1991b, Proc. Natl. Acad. Sci. USA. 88: 8322-8326). The molecular cloning of the cDNA encoding for the PDEB represents the first step in establishing whether this gene plays a causative role in any one of the several human hereditary retinopathies or, based on its localization to chromosome 4p 16.3, in the pathogenesis of Huntington disease.
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PMID:The human beta-subunit of rod photoreceptor cGMP phosphodiesterase: complete retinal cDNA sequence and evidence for expression in brain. 132 54

Hormonal activation of the cGMP-inhibited low Km cyclic AMP phosphodiesterase isoenzyme (cGI.PDE) by effectors, acting either through the cAMP-independent (insulin) or through cAMP-dependent (isoproterenol, forskolin ACTH and 8Br-cAMP) mechanisms, were compared in parametrial (PM) and femoral subcutaneous (SC) adipocytes from sham-operated (SHAM) and ovariectomized (OVX) rats. In SHAM rats, the basal cGI.PDE activity was 50% higher in PM than in SC adipocytes. In OVX rats, the cGi.PDE activatory responses to all the effectors tested remained unchanged in SC, but were completely suppressed in PM adipocytes. The mechanism underlying these defective cGI.PDE activatory responses to cAMP-dependent effectors observed in PM adipocytes after OVX seems to involve protein kinase A, since a decreased activation of cGI.PDE by protein kinase A was also found in these cells. Treatment of OVX rats with both estradiol and progesterone reversed the defective cAMP-dependent activation of cGI.PDE, but not the refractoriness of this isoenzyme to insulin activation. Taken together with previous observations from this laboratory on the fat cell adenylate cyclase system (Lacasa et al. (1991) Endocrinology 128, 747-753), these results: (a) demonstrate that the influence of the ovarian status on the key enzymes controlling cAMP metabolism in fat cells depends on the anatomical origin of these cells, and; (b) provide a biochemical explanation to the insensitivity of the SC adipocyte lipolytic system to ovarian hormones.
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PMID:Hormonal activation of the cGMP-inhibited low-Km cyclic AMP phosphodiesterase of rat adipocytes from different sites: influence of ovariectomy. 132 10

Irish setter dogs affected with rod-cone dysplasia 1 have elevated levels of retinal cGMP resulting from deficient rod-specific cGMP phosphodiesterase (cGMP PDE) activity. We investigated the mRNAs coding for the three subunits of cGMP PDE and for the proteins involved in the activation/deactivation of this enzyme in the retinas of developing affected and control dogs. While the photoreceptor cells are viable in the diseased retinas, opsin, transducin alpha 1 and beta 1, 48 and 33 kd proteins, and cGMP PDE alpha and gamma mRNAs have normal transcript sizes and levels. In contrast, a different pattern of cGMP PDE beta mRNAs with lower than normal concentrations is present in the developing affected retinas prior to degeneration. Our observations suggest that an abnormality involving cGMP PDE beta expression is implicated in rod-cone dysplasia 1.
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PMID:The beta subunit of cyclic GMP phosphodiesterase mRNA is deficient in canine rod-cone dysplasia 1. 132 14

1. The effect of amrinone, milrinone and of three milrinone analogues was tested on spontaneous chronotropic and inotropic activity of guinea-pig isolated atria, on the activity of cGMP-inhibited phosphodiesterase (cGI-PDE) from guinea-pig heart and on specific binding of N6-cyclohexyl[3H]adenosine ([3H]CHA) to Ri adenosine receptors in guinea-pig atria. 2. The Ki-values towards [3H]CHA binding to Ri receptors were linearly related to the EC50S for the increase in force of contraction but not to the EC50S for the increase in frequency of the atria. The Ki values towards cGI-PDE were linearly related to the EC50S for the positive chronotropic effect.
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PMID:Antagonism towards endogenous adenosine and inhibition of cGI-PDE in the cardiac effects of amrinone, milrinone and related analogues. 132 70

Foetal calf serum (FCS) and platelet-derived growth factor (PDGF)-stimulated incorporation of [3H]thymidine into pig aortic smooth muscle cell (ASMC) DNA was decreased by agents that either stimulated the synthesis (forskolin) or inhibited the breakdown (3-isobutyl-1-methylxanthine, IBMX) of cAMP. FCS-stimulated incorporation of [3H]thymidine into DNA was also reduced by selective inhibitors of cAMP-specific phosphodiesterase (PDE IV) (Ro-20-1724, rolipram) and cGMP-inhibited cAMP PDE (PDE III) (SK&F 94836). IBMX, Ro-20-1724, rolipram and SK&F 94836 enhanced forskolin inhibition of DNA synthesis. Alone, rolipram was a relatively weak inhibitor of FCS-induced ASMC DNA synthesis (IC25 greater than 20 microM); however, in the presence of a threshold concentration of SK&F 94836 (20 microM), the potency of rolipram increased (IC25 = 4 microM), suggesting synergy in the actions of PDE III and PDE IV inhibitors. SK&F 94836 and rolipram elicited 30% and 37%, respectively, reductions in FCS-induced ASMC proliferation and potentiated the inhibitory actions of forskolin. PDE III and PDE IV inhibitors alone, exerted minimal effects on ASMC cAMP levels after a short term (10 min) or long-term (2 or 24 hr) exposure, but enhanced forskolin-induced accumulation of cAMP. ASMC spontaneously released cAMP into the extracellular medium, a process that was increased by forskolin. PDE III and PDE IV inhibitors had no effect alone on cAMP extrusion but enhanced the effect of forskolin. Exposure of ASMC to forskolin or SK&F 94836 for 15 min increased the activity ratio (AR) of cAMP-dependent protein kinase from 0.05 to 0.17 and 0.23, respectively. Ro-20-1724, alone, did not affect cAMP-dependent protein kinase but enhanced the stimulatory effect of forskolin (AR = 0.37) and SK&F 94836 (AR = 0.27). Agents that increased cGMP synthesis (glycerol trinitrate, atrial natriuretic factor) or decreased its hydrolysis by selectively inhibiting cGMP-specific PDE (PDE V) (zaprinast) exerted no effects on FCS- or PDGF-stimulated [3H]thymidine incorporation into DNA either alone or in combination. The cytosolic fraction of pig ASMC contained four cyclic nucleotide PDEs which were categorized as PDE V, Ca2+/calmodulin-stimulated PDE (PDE I), PDE III and PDE IV. PDE I and III activities were also associated with the particulate fraction. The results demonstrate that inhibitors of PDEs III and IV alone or in combination with forskolin, reduce ASMC DNA synthesis and proliferation, through an action likely to involve elevation of intracellular cAMP. In contrast, inhibition of cGMP hydrolysing PDE subtypes (I and V) exerted no effect on DNA synthesis in this cell type.
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PMID:Inhibition of pig aortic smooth muscle cell DNA synthesis by selective type III and type IV cyclic AMP phosphodiesterase inhibitors. 132 64


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