Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.12 (PKG)
 2,515 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Dehydroepiandrosterone (DHEA) treatment is effective in preventing or delaying the onset of various genetic and induced disorders of mice and rats. Associated with the beneficial therapeutic effects exerted by action of this steroid is the development of hepatomegaly. To determine whether the changes associated with hepatomegaly also involve alterations in activities of tissue enzymes, we evaluated the effects of DHEA (0.45% in food, w/w) on hepatic protein kinases, phosphatases, and lipogenic enzymes in mice of various strains. The rates of fatty acid and cholesterol syntheses also were evaluated. DHEA administration resulted in profound changes in the sodium dodecylsulfate-polyacrylamide gel electrophoresis patterns of endogenous radiophosphorylated proteins obtained by incubation of liver homogenates with (gamma-32P]ATP. These changes were dependent upon the medium used for homogenization. Thus, when homogenates of liver tissue of DHEA-treated mice were prepared in Tris buffer containing sucrose (0.25 M) there was a marked decrease in phosphorylation of the proteins of relative molecular weight approximately 116,000 (Mr approximately 116,000), approximately 82,000, approximately 80,000, approximately 58,000, approximately 56,000, approximately 48,000, approximately 34,000, and approximately 31,000 compared with controls. With liver homogenates of DHEA-treated mice prepared in Tris buffer alone, there was a marked increase in phosphorylation of the proteins of Mr approximately 70,000, approximately 49,000, approximately 34,000, approximately 31,000, and 28,000 compared with controls. Moreover, the specific activity of kinases for endogenous protein acceptors in liver of control mice was higher than that in liver of DHEA-treated animals. The specific activities of casein kinase, cAMP-dependent protein kinase, and cGMP-dependent protein kinase remained unchanged with DHEA treatment, but the specific activity of histone kinase was increased approximately 30%. Long-term administration of DHEA also was associated with increases in the specific activities of liver AMPase and GTPase (approximately two times), but not of other nucleotidases, alkaline phosphatase, acid phosphatase, glucose-6-phosphatase, or phosphotyrosine phosphatase. The activity of hepatic NADP-linked malic enzyme was increased significantly (two to three times) by DHEA treatment of female mice of three different strains, but was unchanged in male C57BL/6 mice. The specific activities of hepatic glucose-6-phosphate dehydrogenase, NADP-linked isocitrate dehydrogenase, and ATP-citrate lyase were not affected significantly by DHEA treatment of mice. The rate of hepatic lipogenesis, determined by incorporation of tritium from 3H2O into fatty acids, was decreased approximately 70% in DHEA-treated mice, while the rate of cholesterol synthesis was increased approximately 44% compared with controls.
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PMID:Dehydroepiandrosterone feeding and protein phosphorylation, phosphatases, and lipogenic enzymes in mouse liver. 215 82

cAMP-dependent protein kinase, protein kinase C, cGMP-dependent protein kinase, smooth muscle myosin light-chain kinase, and phosphorylase kinase were examined with respect to their ability to phosphorylate porcine atrial muscarinic receptors (mAcChRs). Experiments were performed both in detergent solution and in a reconstituted system containing the mAcChR alone or in the presence of the purified porcine atrial inhibitor guanine nucleotide binding protein (Gi). Only cAMP-dependent protein kinase was capable of phosphorylating the receptor under any of the experimental conditions examined. Phosphorylation of the mAcChR in the detergent-solubilized state resulted in a loss of ligand binding sites that was reversible upon treatment with calcineurin in the presence of calcium and calmodulin. Upon reconstitution, the apparent stoichiometry of phosphorylation was increased by about 15-fold. Carbachol-stimulated covalent incorporation of phosphate was found only in the reconstituted system in the presence of Gi, suggesting that the large agonist-stimulated increase in phosphorylation observed in vivo [Kwatra, M. M., & Hosey, M. M. (1986) J. Biol. Chem. 261, 12429-12432] may in part result from a unique receptor conformation that occurs upon association with this protein. Ligand binding studies indicated that phosphorylation of the mAcChR in the detergent-solubilized or reconstituted state did not affect its interaction with carbachol or L-quinuclidinyl benzilate in vitro. Carbachol-induced stimulation of the GTPase activity of Gi in the reconstituted system was also unaffected by phosphorylation.
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PMID:Phosphorylation of the porcine atrial muscarinic acetylcholine receptor by cyclic AMP dependent protein kinase. 344 51

Nitric oxide (NO) is a potent vasodilator and inhibitor of platelet activation. NO stimulates production of cGMP and activates cGMP-dependent protein kinase (G kinase), which by an unknown mechanism leads to inhibition of Galphaq-phospholipase C-inositol 1, 4,5-triphosphate signaling and intracellular calcium mobilization for several important agonists, including thromboxane A2 (TXA2). To explore the mechanism of platelet inhibition by NO, activation of platelet TXA2 receptors in the presence of cGMP was studied. The nonhydrolyzable analog 8-bromo-cyclic GMP (8-Br-cGMP) potently inhibited activation of the TXA2-specific GTPase in platelet membranes in a concentration-dependent fashion, suggesting that G kinase catalyzes the phosphorylation of some proximal component of the receptor-G protein signaling pathway. Nanomolar concentrations of G kinase were found to catalyze the phosphorylation of platelet TXA2 receptors in vitro, but not Galphaq copurifying with the TXA2 receptors in these experiments. Using immunoaffinity methods, in vivo phosphorylation of TXA2 receptors by cyclic GMP was demonstrated from 32P-labeled cells treated with 8-Br-cGMP. Peptide mapping studies of in vivo phosphorylated TXA2 receptors demonstrated cGMP mediates phosphorylation of the carboxyl terminus of the TXA2 receptor. G kinase also catalyzed the phosphorylation of peptides corresponding to the cytoplasmic tails of both alpha and beta forms of the receptor but not control peptide or a peptide corresponding to the third intracytoplasmic loop of the TXA2 receptor. These data identify TXA2 receptors as cGMP-dependent protein kinase substrates and support a novel mechanism for the inhibition of cell function by NO in which activation of G kinase inhibits signaling by G protein-coupled receptors by catalyzing their phosphorylation.
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PMID:Mechanism of platelet inhibition by nitric oxide: in vivo phosphorylation of thromboxane receptor by cyclic GMP-dependent protein kinase. 956 Jan 98

The septins are a family of GTPase enzymes, some of which are required for the cytokinesis stage of cell division and others of which are associated with exocytosis. We purified and cloned the cDNA for a 40-kDa protein from rat brain that is a substrate for type I cGMP-dependent protein kinase (PKG). The amino acid sequences of two tryptic peptides of P40 showed high homology to the septins. Molecular cloning revealed the 358-amino acid P40 to be a new member of the septin family. P40 was named G-septin, as it is phosphorylated in vitro by PKG, but relatively poorly by the related cAMP-dependent protein kinase and not by protein kinase C. Two splice variants of G-septin (alpha and beta) were found with distinct N and C termini, but a common GTPase domain. G-septin lacks the C-terminal coiled-coil domain characteristic of all other mammalian septins and uniquely has two predicted phosphorylation site motifs for type I PKG. Photoaffinity labeling with [alpha-(32)P]GTP confirmed that G-septin is a GTP-binding protein. Northern blotting showed that G-septin mRNA (5.0 kilobases) is highly expressed in brain and undetectable in 12 other tissues, indicating that the G-septins are primarily neuronal proteins. Very low levels of 6.0-, 3.4-, and 2.6-kilobase transcripts were found in testis. Our results reveal a new class of brain-specific septins that may be regulated by PKG in neurons.
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PMID:Phosphorylation of a new brain-specific septin, G-septin, by cGMP-dependent protein kinase. 1074 83

Small GTPase proteins such as Ras are key regulators of cellular proliferation and are activated by guanine nucleotide exchange/releasing factors (GEFs/GRFs). Three classes of Ras GRFs have been identified to date, represented by Sos1/2, Ras-GRF1/2 and Ras-GRP. Here, we describe a novel candidate Ras activator, cyclic nucleotide rasGEF (CNrasGEF), which contains CDC25, Ras exchange motif (REM), Ras-association (RA), PDZ and cNMP (cAMP/cGMP) binding (cNMP-BD) domains, two PY motifs and a carboxy-terminal SxV sequence. CNrasGEF can activate Ras in vitro, and it binds cAMP directly via its cNMP-BD. In cells, CNrasGEF activates Ras in response to elevation of intracellular cAMP or cGMP, or treatment with their analogues 8-Br-cAMP or 8-Br-cGMP, independently of protein kinases A and G (PKA and PKG). This activation is prevented in CNrasGEF lacking its CDC25 domain or cNMP-BD. CNrasGEF can also activate the small GTPase Rap1 in cells, but this activation is constitutive and independent of cAMP. CNrasGEF is expressed mainly in the brain and is localized at the plasma membrane, a localization dependent on the presence of intact PDZ domain but not the SxV sequence. These results suggest that CNrasGEF may directly connect cAMP-generating pathways or cGMP-generating pathways to Ras.
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PMID:The guanine nucleotide exchange factor CNrasGEF activates ras in response to cAMP and cGMP. 1080 46

Small GTPase Rho and cGMP/cGMP-dependent protein kinase (cGK) pathways exert opposing effects in specific systems such as vascular contraction and growth. However, the direct interaction between these pathways has remained elusive. We demonstrate that cGK phosphorylates RhoA in vitro at Ser188, the same residue phosphorylated by cAMP-dependent protein kinase. In HeLa cells transfected with constitutively active cGK (C-cGK), stress fiber formation induced by lysophosphatidic acid or V14RhoA was blocked. By contrast, C-cGK failed to inhibit stress fiber formation in cells transfected with mutant RhoA with substitution of Ser188 to Ala. C-cGK did not affect actin reorganization induced by Rac1 or Rho-associated kinase, one of the effectors for RhoA. Furthermore, C-cGK expression inhibited the membrane translocation of RhoA. Collectively, our findings suggest that cGK phosphorylates RhoA at Ser188 and inactivates RhoA signaling. The physiological relevance of the direct interaction between RhoA and cGK awaits further investigation.
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PMID:cGMP-dependent protein kinase phosphorylates and inactivates RhoA. 1116 91

Exocytotic secretion is promoted by the concerted action of calcium, guanine nucleotide, and protein kinase C. We now show that the calcium-dependent membrane fusion activity of annexin 7 in vitro is further potentiated by the combined addition of guanine nucleotide and protein kinase C. The observed increment involves the simultaneous activation of annexin 7 by these two effectors. Guanosine triphosphate (GTP) and its non-hydrolyzable analogues optimally enhance the phosphorylation of annexin 7 by protein kinase C in vitro. Reciprocally, phosphorylation by protein kinase C significantly potentiates the binding and hydrolysis of GTP by annexin 7. Only protein kinase C-dependent phosphorylation has a significant positive effect on annexin 7 GTPase, although other protein kinases, including cAMP-dependent protein kinase, cGMP-dependent protein kinase, and pp60(c-)(src), have been shown to label the protein with high efficiency. In vivo, the ratio of bound GDP/GTP and phosphorylation of annexin 7 change in direct proportion to the extent of catecholamine release from chromaffin cells in response to stimulation by carbachol, or to inhibition by various protein kinase C inhibitors. These results thus lead us to hypothesize that annexin 7 may serve as a common site of action for calcium, guanine nucleotide, and protein kinase C in the exocytotic membrane fusion process in chromaffin cells.
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PMID:Protein kinase C and guanosine triphosphate combine to potentiate calcium-dependent membrane fusion driven by annexin 7. 1199 95

The slow EPSP (sEPSP) or slow EPSC (sEPSC) at parallel fiber to Purkinje neuron synapses is attributable to a nonselective cation channel coupled to activation of metabotropic type 1 glutamate receptors (mGluR1s). Photorelease of L-glutamate in 1 msec from 4-methoxy-7-nitroindolinyl-or 7-nitroindolinyl-caged glutamate in cerebellar slices was used to isolate and study postsynaptic mechanisms coupling mGluR1 to the cation channel. L-Glutamate immediately activated a glutamate transporter current, followed by the slow mGluR1-activated conductance. Inhibitors of kinases, phosphatases, and G-proteins were tested on the peak glutamate-evoked currents. No effects of the inhibitors were seen on the initial glutamate transporter currents. In contrast, the later mGluR1 currents were either unaffected or enhanced by the protein tyrosine kinase (PTK) inhibitors PP1, K252a, and staurosporine were diminished or blocked by phosphatase inhibitors but were unaffected by inhibitors of serine-threonine kinases PKA, PKC, or PKG. The selective src-PTK inhibitor PP1 (10 microm intracellularly) potentiated submaximal mGluR1 currents evoked by low L-glutamate concentrations but had no effect on maximal responses (80 or 160 microm L-glutamate). L-Glutamate-evoked mGluR1 currents and parallel fiber sEPSCs were reversibly and completely inhibited by protein tyrosine phosphatase (PTP) inhibitor bpV(phen) (50-200 microm) and by nonselective phosphatase inhibitor orthovanadate (0.5 or 1 mm). mGluR1 currents were completely inhibited by GDPbetaS applied intracellularly (5 mm). The results confirm a role for a GTPase postsynaptically, show that tyrosine phosphorylation inhibits mGluR1 coupling to the channel, and show that PTPs increase activation by tyrosine dephosphorylation most likely upstream of the sEPSP cation channel.
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PMID:Evidence for protein tyrosine phosphatase, tyrosine kinase, and G-protein regulation of the parallel fiber metabotropic slow EPSC of rat cerebellar Purkinje neurons. 1276 93

Nitric oxide (NO) inhibits vascular contraction by activating cGMP-dependent protein kinase I-alpha (PKGI-alpha), which causes dephosphorylation of myosin light chain (MLC) and vascular smooth muscle relaxation. Here we show that PKGI-alpha attenuates signaling by the thrombin receptor protease-activated receptor-1 (PAR-1) through direct activation of regulator of G-protein signaling-2 (RGS-2). NO donors and cGMP cause cGMP-mediated inhibition of PAR-1 and membrane localization of RGS-2. PKGI-alpha binds directly to and phosphorylates RGS-2, which significantly increases GTPase activity of G(q), terminating PAR-1 signaling. Disruption of the RGS-2-PKGI-alpha interaction reverses inhibition of PAR-1 signaling by nitrovasodilators and cGMP. Rgs2-/- mice develop marked hypertension, and their blood vessels show enhanced contraction and decreased cGMP-mediated relaxation. Thus, PKGI-alpha binds to, phosphorylates and activates RGS-2, attenuating receptor-mediated vascular contraction. Our study shows that RGS-2 is required for normal vascular function and blood pressure and is a new drug development target for hypertension.
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PMID:Regulator of G-protein signaling-2 mediates vascular smooth muscle relaxation and blood pressure. 1460 79

The septins are a family of GTPase enzymes required for cytokinesis and play a role in exocytosis. Among the ten vertebrate septins, Sept5 (CDCrel-1) and Sept3 (G-septin) are primarily concentrated in the brain, wherein Sept3 is a substrate for PKG-I (cGMP-dependent protein kinase-I) in nerve terminals. There are two motifs for potential PKG-I phosphorylation in Sept3, Thr-55 and Ser-91, but phosphoamino acid analysis revealed that the primary site is a serine. Derivatization of phosphoserine to S-propylcysteine followed by N-terminal sequence analysis revealed Ser-91 as a major phosphorylation site. Tandem MS revealed a single phosphorylation site at Ser-91. Substitution of Ser-91 with Ala in a synthetic peptide abolished phosphorylation. Mutation of Ser-91 to Ala in recombinant Sept3 also abolished PKG phosphorylation, confirming that Ser-91 is the major site in vitro. Antibodies raised against a peptide containing phospho-Ser-91 detected phospho-Sept3 only in the cytosol of nerve terminals, whereas Sept3 was located in a peripheral membrane extract. Therefore Sept3 is phosphorylated on Ser-91 in nerve terminals and its phosphorylation may contribute to the regulation of its subcellular localization in neurons.
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PMID:Phosphorylation of septin 3 on Ser-91 by cGMP-dependent protein kinase-I in nerve terminals. 1510 17


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