Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mutations in the lon (capR) gene result in multiple phenotypes, one of which is the failure to degrade abnormal and normal proteins (Deg-). Previous work with partially purified preparations showed that the lon (capR) gene product is a 94,000-dalton polypeptide with an affinity for nucleic acids. The lon (capR) protein has now been highly purified and is demonstrated to have an ATP-dependent protease activity. The enzyme hydrolyzed 3H-labeled alpha-casein into trichloroacetic acid-soluble forms in Tris buffer containing Mg2+ and ATP. The reaction has a pH optimum of 8.5 and ATP was the preferred nucleotide. CTP and UTP could substitute for ATP (75% and 67%, respectively) but GTP, ADP, AMP, cyclic AMP, and PPi could not. Proteolysis by the lon (capR) protein required ATP hydrolysis. Nonhydrolyzable analogs of ATP and CTP did not promote casein cleavage. When low concentrations of ATP were used, proteolysis stopped as the ATP pool was depleted. Casein stimulated lon (capR) ATPase activity, and the products were ADP and inorganic phosphate in equimolar amounts. No protein kinase activity was detected. The DNA-binding activity, present in partially pure preparations, was retained in the purified protein. The gene product purified from a lon nonsense mutant that exhibits the Deg- phenotype (capR9), lacked both the ATP-dependent protease and ATPase activities, though it retained DNA-binding activity. Absence of an ATP-dependent protease activity could account for many of the pleiotropic effects observed in lon mutants.
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PMID:ATP hydrolysis-dependent protease activity of the lon (capR) protein of Escherichia coli K-12. 645 36

The middle T antigen (MT Ag) encoded by polyoma virus has an associated protein kinase activity which transfers a phosphoryl group from ATP or GTP to a tyrosine residue on MT Ag in immunoprecipitates formed between polyoma virus-infected or transformed cell extracts and serum from animals bearing polyoma-induced tumors. Incubation of such immunoprecipitates or polyoma-transformed cell extracts prior to immunoprecipitation with the sulfhydryl reagent, N-ethylmaleimide (NEM), resulted in a significant inhibition of MT Ag-associated kinase activity. Inactivation of this enzyme activity by NEM was found to be dependent upon the incubation pH, time of incubation, and NEM concentration. ATP, GTP, and ADP in the presence of Mg2+ were found to decrease the rate of NEM-mediated inactivation of MT Ag-associated kinase activity, while CTP and UTP did not detectably alter the rate of enzyme inhibition by NEM. These results suggest that the MT Ag-associated kinase possesses at least one NEM-sensitive sulfhydryl group essential for phosphotransferase activity which may be present at or near the enzyme catalytic site.
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PMID:Inhibition of polyoma virus middle T antigen-associated tyrosyl kinase activity by N-ethylmaleimide. 655 75

Several characteristics of receptor capping in lymphocyte membranes suggest similarities with mechanisms underlying control of contraction in smooth muscle fibers. Both capping and contraction are Ca2+ dependent and require metabolic energy. Contractile proteins such as actin and myosin are associated with the cap, as is calmodulin, which mediates the Ca2+ dependence of smooth muscle contraction. Recent studies have shown that myosin light chain kinase (MLCK), which plays a central role in regulation of smooth muscle contraction, is also present in isolated lymphocyte membrane-cytoskeleton complexes. We have explored this analogy further, using mouse lymphoma T cells whose membranes were rendered permeable to small proteins by using a low-Ca2+ EGTA solution similar to that used to chemically skin smooth muscle cells. Permeabilized lymphocytes were then exposed to solutions containing various combinations of high or low Ca2+, ATP, or other nucleotides (5'-adenylyl imidodiphosphate, adenosine 5'-[gamma-thio]triphosphate, guanosine 5'-[gamma-thio]triphosphate, CTP, ITP, UTP, and GTP), calmodulin, Ca2+-insensitive MLCK (MLCK subunit that has been stripped of the Ca2+ binding site), and the catalytic subunit of cAMP-dependent protein kinase that phosphorylates (and thereby inactivates) MLCK. Capping of concanavalin A-labeled receptors in these various test solutions was scored. In all solutions the capping observed in permeable lymphoma cells correlated well with contraction previously observed in similarly treated skinned smooth muscle fibers, providing strong evidence for the involvement of myosin light chain phosphorylation in the regulation of receptor capping.
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PMID:Regulation of receptor capping in mouse lymphoma T cells by Ca2+-activated myosin light chain kinase. 658 74

In the present study, we examined the effects of guanine nucleotides on vasopressin-induced osmotic water flow and sodium transport in the 14-h preincubated frog bladder. We also examined the effects of the adenylate cyclase-cyclic AMP and cyclic AMP-dependent protein kinase system in the bladder's epithelial cells. Gpp(NH)p significantly enhanced vasopressin-induced water flow while it did not affect cyclic AMP-induced water flow. However, Gpp(NH)p did not enhance the vasopressin-induced increment of sodium transport across the frog bladder. The adenylate cyclase activity of the crude homogenate was enhanced by vasopressin, Gpp(NH)p and NaF. The effects of Gpp(NH)p and vasopressin, at their maximum doses, on the enzyme activities were additive, while other combinations were not. Specific Gpp(NH)p binding sites were found in the pellet fraction after 2,400 X g centrifugation. No direct effect on the protein kinase activity was observed in the presence of 10(-6) M nucleotides, such as GTP, GDP, GMP, CTP, UTP, ITP and Gpp(NH)p. Cyclic AMP stimulated the phosphorylation of discrete protein bands, however, Gpp(NH)p did not influence cyclic AMP-dependent protein phosphorylation of crude homogenate of the bladder's epithelial cells. These results suggest the guanine nucleotides stimulate the vasopressin-induced osmotic water flow in frog bladder by enhancing the vasopressin-mediated adenylate cyclase activity, so that accumulated cyclic AMP might activate cyclic AMP-dependent protein kinase.
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PMID:Effects of guanine nucleotides on vasopressin-induced water flow and sodium transport of the frog bladder. 660 7

Uptakes of radioactive C1- or 1- by gastric microsomal vesicles were stimulated 2- to 8-fold by AtP. The sensitivity of those uptakes to a C1- in equilibrium OH- ionophore and to osmotic swelling suggested they were due to transport rather than to binding. The ATP effect was labile, but dithiothreitol and methanol improved its stability. The stimulation of anion transport required magnesium; GTP and UTP were less potent than ATP whereas ADP and AMP had no effect. The apparent Km for ATP was estimated to be 2 X 10(-4) M at 22 degrees C. The rate of the ATP-dependent transport showed saturation-type kinetics, with half-maximal uptake at 10 mM for I- and 15 mM for C1-. Nonradioactive C1-, I-, and SCN- competed with 125I- uptake while SO42- did not. K+ valinomycin increased the ATP-dependent C1- uptake. The thermostable inhibitor of cAMP-dependent protein kinases inhibited the effect of ATP. These results suggest the existence of an anion conductance, permeant to C1-, I-, and SCN- and nonpermeant to SO42-, which could be linked to a cAMP-dependent protein kinase.
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PMID:C1-transport in gastric micorsomes. An ATP-dependent influx sensitive to membrane potential and to protein kinase inhibitor. 744 May 65

ATP produced whole-cell potassium currents in cultured endothelial cells of the bovine brain cortical arteries. P2 purinoceptor agonists evoked similar currents with the order of their potency: 2-methylthio ATP > ATP >> alpha, beta-methylene ATP > or = UTP > or = ADP >> AMP. ATP-evoked currents were inhibited by GDP beta S, but not by pertussis toxin (PTX). Furthermore, a phospholipase C (PLC) inhibitor, protein kinase C inhibitor, or cAMP-dependent protein kinase inhibitor had no effect on the currents. In addition to these effects, ATP enhanced intracellular free Ca2+ concentration ([Ca2+]i) in the presence and absence of extracellular Ca2+, and this [Ca2+]i increase was not inhibited by a PLC inhibitor. These results, thus, provide an indication that ATP activates the potassium channel and enhances [Ca2+]i via a P2Y purinoceptor linked to a PTX-insensitive G-protein, which is not involved in a PLC-mediated signaling pathway.
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PMID:ATP activates the potassium channel and enhances cytosolic Ca2+ release via a P2Y purinoceptor linked to pertussis toxin-insensitive G-protein in brain artery endothelial cells. 748 26

Basolateral Na-K-Cl cotransport activity in primary cultures of dog tracheal epithelial cells is stimulated by beta-adrenergic agents, such as isoproterenol, and by apical UTP, which acts through an apical P2-purinergic receptor. While at least part of the stimulatory effect of isoproterenol appears to involve direct activation of the cotransporter via cAMP-dependent protein kinase, cotransport stimulation by apical UTP is entirely secondary to apical Cl- efflux and a resultant decrease in intracellular [Cl-] ([Cl-]i) and/or cell shrinkage (Haas, M., and McBrayer, D. G. (1994) Am. J. Physiol. 266, C1440-C1452). In the secretory epithelia of the shark rectal gland and avian salt gland, Na-K-Cl cotransport activation by both cAMP-dependent and cAMP-independent secretagogues has been shown to be accompanied by phosphorylation of the cotransport protein itself (Lytle, C., and Forbush, B., III (1992) J. Biol. Chem. 267, 25438-25443; Torchia, J., Lytle, C., Pon, D. J., Forbush, B., III, and Sen, A. K. (1992) J. Biol. Chem. 267, 25444-25450). In the present study, we immunoprecipitate the approximately 170-kDa Na-K-Cl cotransport protein of dog tracheal epithelial cells with a monoclonal antibody against the cotransporter of the intestinal cell line T84. Incubation of confluent primary cultures of tracheal cells with isoproterenol and apical UTP increases basolateral-to-apical 36Cl- flux 3.4- and 2.6-fold, respectively, and produces similar increases (3.2- and 2.8-fold, respectively) in 32P incorporation into the approximately 170-kDa cotransport protein. Decreasing [Cl-]i (without concomitant cell shrinkage) by incubating cultures with apical nystatin and reduced apical [Cl-] ([Cl-]alpha) likewise increases both cotransport activity and cotransport protein phosphorylation. These effects become more pronounced with greater reductions in [Cl-]alpha; after 20 min of incubation with nystatin and 32 mM [Cl-]alpha, cotransport activity and 32P incorporation into the cotransport protein are increased 2.8- and 2.7-fold, respectively, similar to increases seen with apical UTP. 2-3-fold increases in cotransporter activity and phosphorylation are also seen in nystatin-treated cells under hypertonic conditions (50 mM sucrose added apically and basolaterally). These findings suggest a close correlation between Na-K-Cl cotransport activity and phosphorylation of the approximately 170-kDa cotransport protein. The latter is phosphorylated in response to both reduced [Cl-]i and cell shrinkage, either or both of which are likely to be involved in secondary cotransport activation in response to apical UTP.
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PMID:[Cl-]i-dependent phosphorylation of the Na-K-Cl cotransport protein of dog tracheal epithelial cells. 749 26

The cystic fibrosis transmembrane conductance regulator (CFTR) is a Cl- channel activated by protein kinase A and regulated by ATP in a complex manner. We have applied patch-clamp techniques to C127i mouse mammary carcinoma cells transfected with human CFTR to assess the role of external ATP in the modulation of CFTR function. Extracellular ATP was sufficient to activate non-rectifying, Cl(-)-selective whole-cell currents in CFTR-transfected, but not mock-transfected cells. The ATP-mediated activation of CFTR was independent of protein kinase A since channel activation by ATP was preserved in cells that were (a) depleted of intracellular ATP, (b) incubated with the cAMP antagonist Rp-cAMPS, or (c) exposed to the protein kinase A inhibitor, 5-24 amide. In each of these conditions, 8-Br-cAMP was no longer capable of activating CFTR. The possibility that the extracellular ATP activation of Cl- currents in CFTR-expressing C127i cells was mediated by a P2-type purinergic receptor was supported by studies in which the effect of external ATP on the Cl- currents was mimicked by the ATP analogs, ATP gamma S and beta,gamma-methylene ATP, but not the uridine nucleotide, UTP. Single-channel analysis of ATP-activated Cl -currents under both cell-attached and excised, inside-out patch-clamp configurations indicated that this channel is only present in CFTR-transfected cells and indistinguishable from CFTR. External ATP also activated ATP currents in CFTR-transfected cells, a novel function of CFTR. These findings are consistent with the presence of a purinergic receptor signal transduction mechanism in C127i cells whose activation by external ATP is linked to the activation of CFTR in a cAMP-independent manner. The data provide additional support for the use of ATP and its analogs as alternative therapies in cystic fibrosis.
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PMID:External ATP and its analogs activate the cystic fibrosis transmembrane conductance regulator by a cyclic AMP-independent mechanism. 751 60

Several authors have shown that angiotensin II stimulates hepatic angiotensinogen synthesis in vivo, ex vivo and in vitro. In previous studies we have demonstrated that this effect of angiotensin II depends mainly on a transient inhibition of adenylyl cyclase and is the consequence of a stabilization of angiotensinogen mRNA. In the present study we describe the isolation of a polysomal 12 kDa protein which, in band shift and cross link assays, shows a specific affinity to the 3' untranslated region (3' UTR) of angiotensinogen mRNA and prevents enzymatic degradation of angiotensinogen mRNA in a cell-free incubation system. [32P]UTP-labelled or unlabelled 3' fragments of angiotensinogen mRNA were synthesized on a transcription vector (pGEM5zf+) into which the corresponding DNA sequence was cloned after restriction from vector pRAG 16. Binding of the 12 kDa protein to the radioactively labelled 3' UTR of angiotensinogen mRNA could be displaced by unlabelled 3' UTR mRNA fragments but not by a renin mRNA of comparable length derived from the coding region. The RNA-binding protein appears to be derived from a higher molecular mass precursor (45 kDa) which is preferentially present under reducing conditions in vitro; the active low molecular mass form is evident in the absence of reducing agents. In a cross link experiment we established that a band shift signal which was obtained in the presence of the 45 kDa protein preparation exclusively depends on RNA binding of the active 12 kDa protein. In addition, a phosphorylation step may be involved in the activation of the 12 kDa protein, since its molecular mass and isoelectric point correlate with proteins which were phosphorylated in response to transient decreases of cAMP (induced by guanfacine or angiotensin II) or in response to a direct inhibition of protein kinase A by the cAMP antagonist Rp-cAMP. The importance of phosphorylation reactions for the stabilization of angiotensinogen mRNA was further assessed in a cell-free incubation system of rat liver parenchymal cells. These studies demonstrated that in the presence of acid phosphatase (1 U/ml) the half-life of angiotensinogen was significantly decreased. In the same incubation system the 12 kDa protein increased the half-life of endogenous as well as of exogenous angiotensinogen mRNA three- to fourfold, while no stabilizing effect was apparent when exogenous angiotensinogen mRNA from which the 3' tail had been deleted was added.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Contribution of a 12 kDa protein to the angiotensin II-induced stabilization of angiotensinogen mRNA: interaction with the 3' untranslated mRNA. 761 10

This report demonstrates that platelets possess P2 purinoceptors with unique properties that distinguish them from the ADP (P2T) receptor. Extracellular ATP, and its poorly hydrolyzable analogues, inhibit collagen- and U46619 (a thromboxane mimetic)-induced platelet aggregations. Adenosine deaminase was without effect on ATP action while reversing the inhibitory effect of adenosine. A unique aspect of the P2 receptor is the sensitivity to UTP and CTP and insensitivity to GTP. The rank order of inhibition by beta gamma-methylene ATP, alpha beta-methylene ATP > ATP indicates that a P2x-like receptor is present on the platelet membrane. This conclusion is further supported by the nearly complete desensitization to ATP by pre-exposure of platelets to alpha beta-methylene-ATP. However, unlike previously described P2x purinoceptors, the inhibition of platelet aggregation by extracellular ATP appears to result, at least in part, from the ATP-induced increase of intracellular cyclic AMP levels apparently coupled through a Gs protein. The combined addition of iloprost (0.14 to 1.39 nM) and ATP (18 microM) or ATP (20-40 microM) and the phosphodiesterase inhibitor theophylline (0.5 to 1 mM) synergistically inhibited platelet aggregation implying a common interactive site with adenylate cyclase. This is further substantiated by the ability of the adenylate cyclase inhibitor, 2',5'-dideoxyadenosine, to abrogate the inhibitory effects of ATP. The protein kinase A (PKA) inhibitor H1004 blocks ATP inhibition of platelet aggregation while the protein kinase C inhibitor H7 did not. This implies that the generation of cyclic AMP, with the subsequent activation of PKA and phosphorylation of selected proteins is required, in part, for the action of ATP.
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PMID:Occupancy of P2 purinoceptors with unique properties modulates the function of human platelets. 768 12


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