Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.16.2 (PCP)
 3,761 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The effects of the benzopyran K-channel opener, BRL55834, on mechanical activity in bovine trachealis and rat portal vein were studied together with membrane currents in freshly-isolated single cells derived from these tissues. 2. BRL55834 (3 nM-1 microM) produced a concentration-dependent relaxation of bovine trachealis precontracted with 100 microM histamine and reduced the spontaneous mechanical activity of rat portal veins, effects which were antagonized by glibenclamide (1-10 microM) but were not reversible on washing. In contrast, charybdotoxin (250 nM) did not modify the spasmolytic effect of BRL55834 in bovine trachealis. 3. BRL55834 (10 nM-10 microM) did not relax segments of bovine trachealis precontracted with 80 mM KCl. 4. In some freshly-isolated single cells from bovine trachealis held at -10 mV, BRL55834 (3 microM) induced a time-independent outward K-current which was partially resistant to inhibition by glibenclamide (10 microM). In other cells, a very noisy, outwardly-rectifying and charybdotoxin-sensitive current developed in the presence of BRL55834 (3 microM) and in time-matched control cells. 5. In freshly-isolated single cells from rat portal vein held at -10 mV, BRL55834 (3 microM) induced a time- and calcium-independent outward K-current which was partially resistant (approximately 25% inhibition at +40 mV) to subsequent inhibition by glibenclamide (10 microM). In contrast, levcromakalim induced a time-independent outward K-current which was completely inhibited by glibenclamide 10 microM. 6. With the non-hydrolysable ATP analogue, AMP-PCP (5 mM), in the pipette, the ability of BRL55834 to induce a time-independent K-current in portal vein cells was markedly reduced (approximately 80% inhibition at +40 mV) whereas the effects of 10 microM levcromakalim were totally inhibited. 7. The glibenclamide-resistant current component induced by BRL55834 was totally inhibited by phentolamine (100 microM), a concentration that had no effect on the peak current (IBK(Ca)) induced by NS1619 (33 microM). 8. Stationary fluctuation analysis of the noise associated with the glibenclamide-insensitive K-current induced by BRL55834 in rat portal vein cells indicated that the unitary current flowing through the underlying channels was 0.26 pA at -10 mV, a value inconsistent with the involvement of BKCa. 9. It is concluded that the relaxations of both bovine trachea and rat portal vein produced by BRL55834 are associated with the opening of K-channels. These are probably identical to the ATP-sensitive K-channel opened by levcromakalim, although the involvement of an additional K-channel cannot be excluded. The reduced sensitivity of the BRL55834-induced changes to glibenclamide and toAMP-PCP may result from avid binding of BRL55834 to its site of action.
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PMID:Effects of BRL55834 in rat portal vein and bovine trachea: evidence for the induction of a glibenclamide-resistant, ATP-sensitive potassium current. 758 99

Excess ATP is known to enhance Ca(2+)-ATPase activity and, among other effects, to accelerate the Ca2+ binding reaction. In previous work, we studied the pH dependence of this reaction and proposed a 3H+/2Ca2+ exchange at the transport sites, in agreement with the H+/Ca2+ counter transport. Here we studied the effect of ADP and nonhydrolyzable ATP analogues on the Ca2+ binding reaction at various pH values. At pH 6, where Ca2+ binding is monophasic and slow, ADP, adenosine 5'-(beta,gamma-methylene)triphosphate (AMP-PCP), or adenyl-5'-yl imidodiphosphate (AMPPNP) increased the Ca2+ binding rate constant 20-fold. At pH 7 and 8, where Ca2+ binding is biphasic, the nucleotides induce fast and monophasic Ca2+ binding. At pH 7, AMP-PCP accelerated Ca2+ binding with an apparent dissociation constant of 10 microM. At acidic pH, ADP, AMPPCP, or AMPPNP increased the equilibrium affinity of Ca2+ for ATPase, whereas at alkaline pH, these nucleotides had no effect. At pH 5.5, AMPPCP increased equilibrium Ca2+ binding with an apparent dissociation constant of 1 microM.
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PMID:The modulation of Ca2+ binding to sarcoplasmic reticulum ATPase by ATP analogues is pH-dependent. 759 71

The sympathetic nervous system has been shown to exert a trophic influence on vascular smooth muscle cells (VSMC). Therefore, we studied the growth-regulating effects of the sympathetic cotransmitters ATP, neuropeptide Y (NPY), and norepinephrine (NE). ATP in concentrations of 1-100 microM greatly increased the incorporation of [3H]thymidine in VSMC from rat aorta and vena cava. ATP also increased cell number and total protein content. The maximal effect on [3H]thymidine incorporation was greater than for epidermal growth factor (20 ng/ml) or insulin (1 microgram/ml) and approximately one-half that of 10% fetal calf serum. The potency series of other nucleotides and analogues of ATP was ATP > beta, gamma-methyleneATP (AMP-PCP) > ADP > adenosine > alpha, beta- methyleneATP (AMP-CPP) > 2-methylthioATP, indicating involvement of a P2 receptor, however, it does not meet proposed pharmacological criteria of either the P2x or P2y subclass. Several proposed P2 receptor antagonists were without effect. The effect of ATP could be mediated by a "nucleotide receptor," since UTP also stimulated [3H]thymidine incorporation. In our model, there was a strong correlation between the mitogenic effects of ATP, AMP-CPP, AMP-PCP, and UTP and their ability to stimulate influx of extracellular Ca2+ (Ca2+o). Moreover, the mitogenic effect of ATP was increased by high concentrations of Ca2+o. Taken together with data showing the lack of involvement of several other second-messenger systems, this indicates a critical role for Ca2+o in mediating the mitogenic effects of ATP. Amiloride, known to inhibit the action of several growth factors, also inhibited ATP-induced mitogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Mitogenic effects of ATP on vascular smooth muscle cells vs. other growth factors and sympathetic cotransmitters. 769 83

We have identified a 70-kDa cytosolic protein (GTBP70) in rat adipocytes that binds to glutathione S-transferase fusion proteins corresponding to the cytoplasmic domains of the facilitative glucose transporter isoforms Glut1, Glut2, and Glut4. GTBP70 did not bind to irrelevant fusion proteins, indicating that the binding is specific to the glucose transporter. GTBP70 binding to the glucose transporter showed little isoform specificity but was significantly subdomain-specific; it bound to the C-terminal domain and the central loop, but not to the N-terminal domain of Glut4. The GTBP70 binding to Glut4 was not affected by the presence of 2 mM EDTA, 2.4 mM Ca2+, or 150 mM K+. The binding was inhibited by ATP in a dose-dependent manner, with 50% inhibition at 10 mM ATP. This inhibition was specific to ATP, as ADP and AMP-PCP (adenosine 5'-(beta, gamma-methylenetriphosphate)) were without effect. GTBP70 did not react with antibodies against phosphotyrosine, phosphothreonine, or phosphoserine, suggesting that it is not a phosphoprotein. The binding of GTBP70 to Glut4 was not affected by the pretreatment of adipocytes with insulin. When these experiments were repeated using rat hepatocyte cytosols, no ATP-sensitive 70-kDa protein binding to the glucose transporter fusion proteins was evident, suggesting that either GTBP70 expression or its function is cell-specific. These findings strongly suggest the possibility that GTBP70 may play a key role in glucose transporter regulation in insulin target cells such as adipocytes.
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PMID:ATP-sensitive binding of a 70-kDa cytosolic protein to the glucose transporter in rat adipocytes. 771 80

1. This paper identifies and characterizes an ATP-dependent copper transport system in endoplasmic reticulum vesicles isolated from male rat liver. 2. The transporter has a Km of 2.5 +/- 1.2 mumol 1(-1) copper glutathione (CuGSH) and a Vmax of 4.5 +/- 1.3 nmol (mg protein)-1 (5 min)-1 for copper. 3. At a copper concentration of 2 mumol l-1, ATP dependence reaches saturation, with a Km for ATP of 4.7 +/- 2.4 mmol l-1 and a Vmax of 2.8 +/- 0.6 nmol (mg protein)-1 (5 min)-1. 4. The uptake is dependent on ATP hydrolysis, since a low energy analogue of ATP, adenosine 5'-[beta-gamma-methylene] triphosphate tetralithium (AMP.PCP), has no effect on copper uptake. 5. The transporter is a P-type ATPase, since vanadate inhibits uptake with a high degree of specificity (100 mumol l-1 inhibits uptake by 50% at a copper concentration of 2 mumol l-1).
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PMID:Identification of an ATP-dependent copper transport system in endoplasmic reticulum vesicles isolated from rat liver. 773 49

Yeast mitochondria were found to contain a novel topoisomerase-like activity which required nucleoside di- or tri-phosphates as a cofactor. ADP supported activity as effectively as ATP and the optimal concentration for each was approximately 20 microM. None of the other standard ribo- or deoxyrib-onucleotides could fully substitute for either ADP or ATP. The non-hydrolyzable ATP analogs, adenosine-5'-0-(3-thiotriphosphate) (ATP-gamma-S), adenylyl (beta,gamma-methylene) (AMP-PCP), and andenyl-imidodiphosphate (AMP-PNP) also supported activity suggesting that the nucleotide cofactor regulated topoisomerase activity rather than serving as an energy donor in the reaction. The mitochondrial topoisomerase activity relaxed both positively and negatively supercoiled DNA. It was not inhibited by concentrations of ethidium bromide up to 2 micrograms/ml nor by either nalidixic or oxolinic acids; novobiocin, coumermycin, and berenil inhibited the activity. Genetic and biochemical analysis of the mitochondrial topoisomerase activity indicated that it was not encoded by the nuclear TOP1, TOP2, and TOP3 genes.
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PMID:Evidence for a nucleotide-dependent topoisomerase activity from yeast mitochondria. 775 Jan 44

1. Myotoxin alpha (MYTX), a polypeptide toxin purified from the venom of prairie rattlesnakes (Crotalus viridis viridis) induced Ca2+ release from the heavy fraction (HSR) but not the light fraction of skeletal sarcoplasmic reticulum at concentrations higher than 1 microM, followed by spontaneous Ca2+ reuptake by measuring extravesicular Ca2+ concentrations using the Ca2+ electrode. 2. The rate of 45Ca2+ release from HSR vesicles was markedly accelerated by MYTX in a concentration-dependent manner in the range of concentrations between 30 nM and 10 microM, indicating the most potent Ca2+ releaser in HSR. 3. The Ca2+ dependency of MYTX-induced 45Ca2+ release has a bell-shaped profile but it was quite different from that of caffeine, an inducer of Ca(2+)-induced Ca2+ release. 4. 45Ca2+ release induced by MYTX was remarkable in the range of pCa between 8 and 3, whereas that by caffeine was prominent in the range of pCa, i.e., between 7 and 5.5. 5. MYTX-induced 45Ca2+ release consists of both early and late components. The early component caused by MYTX at low concentrations (30-300 nM) completed within 20 s, while the late component induced by it at higher concentrations (> 0.3 microM) was maintained for at least 1 min. 6. Both the components were almost completely inhibited by inhibitors of Ca2+ such as Mg2+, ruthenium red and spermine. 7. 45Ca2+ release induced by caffeine or beta,gamma-methyleneadenosine 5'-triphosphate (AMP-PCP) was completely inhibited by high concentrations of procaine. Procaine abolished the early component but not the late one, suggesting that at least the early component is mediated through Ca(2+)-induced Ca2+ release channels. 8. On the basis of these results, the character of Ca2+ release induced by MYTX was quite different from that caused by caffeine or AMP-PCP, suggesting that MYTX induces Ca2+ release having novel properties in HSR. MYTX is the first polypeptide Ca2+ inducer and has become a useful pharmacological tool for clarifying the mechanism of Ca2+ release from skeletal muscle SR.
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PMID:Ca2+ release induced by myotoxin alpha, a radio-labellable probe having novel Ca2+ release properties in sarcoplasmic reticulum. 781 16

The regulation of Cl- and cation conductances by the nonhydrolyzable ATP analog adenosine 5'-(beta,gamma-methylene)triphosphate (AMP-PCP) was characterized in isolated zymogen granules (ZG) from pancreatic acinar cells. ZG were purified from rat pancreas homogenate by Percoll gradient centrifugation. Cl- conductance was assayed by suspending ZG in isotonic KCl buffer and measuring osmotic lysis induced by maximal permeabilization of ZG membranes (ZGM) for K+ with the K+ ionophore valinomycin (Val). This resulted in influx of K+ through the artificial pathway and of Cl- through endogenous channels. To measure cation conductances ZG (pHi approximately 6) were suspended in pH 7 buffered isotonic monovalent cation acetate salts. The pH gradient was converted into an outside-directed H+ diffusion potential by maximally increasing H+ conductance of ZGM with the protonophore carbonyl cyanide p-chlorophenylhydrazone. Osmotic lysis of ZG was induced by H+ diffusion potential driven influx of monovalent cations through endogenous channels and non-ionic diffusion of the counterion acetate. In the absence of Val, ZG were stable in KCl buffer up to 2 h. AMP-PCP enhanced osmotic lysis approximately 4-fold compared to control, due to activation of Cl- conductance by AMP-PCP and K+ influx through an AMP-PCP-insensitive nonselective cation pathway, which could be blocked by 0.1 mM Ba2+, 0.5 mM quinine, or 0.2 mM flufenamate. In addition, a K+ and Rb+ selective cation conductance was found which was completely blocked by 0.5 mM AMP-PCP or 0.5 mM quinine. AMP-PCP induced Cl- conductance was strongly inhibited by two monoclonal antibodies against MDR1 P-glycoprotein (JSB-1 and C219; 5-10 micrograms/ml), but not by a monoclonal antibody against the cystic fibrosis transmembrane conductance regulator (M3A7; 5 micrograms/ml) or by mouse IgG. The AMP-PCP insensitive nonselective cation conductance was not blocked by monoclonal antibodies against MDR1 P-glycoprotein (MDR1). Immunoblot studies of ZG membranes revealed the presence of a major immunoreactive protein band of approximately 65 kDa with both monoclonal antibodies against MDR1, but no protein of the approximate size of MDR1 (approximately 170 kDa) was detected. We propose that the Cl- channel or a regulator of the channel, that is activated by the non-hydrolyzable ATP analog AMP-PCP in ZG membranes, is a member of the ATP binding cassette superfamily of transporters and may have homology to MDR1 P-glycoprotein.
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PMID:Monoclonal antibodies against MDR1 P-glycoprotein inhibit chloride conductance and label a 65-kDa protein in pancreatic zymogen granule membranes. 792 2

We have previously demonstrated in permeabilized rat pancreatic acini that the existence of two affinity states of the pancreatic cholecystokinin (CCK) receptor seen in intact cells depends on the presence of ATP. In the present study, we demonstrate that this effect of ATP is mediated by the enzyme nucleoside diphosphate kinase (NDPK). Northern blot hybridization analysis demonstrated NDPK mRNA in pancreas. Furthermore, pancreatic membranes possessed NDPK activity, which transferred high-energy phosphate groups to [8-3H]GDP. This enzyme also utilized UTP and ITP as a source of gamma-phosphate for GTP formation while guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) was formed in the presence of adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S). However, adenylyl (beta, gamma-methylene)-diphosphate (AMP-PCP) did not serve as a substrate for NDPK. Analysis of 125I-Bolton-Hunter-labeled CCK octapeptide ([125I]BH-CCK-8) binding data in the absence of nucleotides was consistent with a single affinity state with dissociation constant (Kd) equal to 80 pM and maximal binding equal to 50.8 fmol/mg. ATP, UTP, ITP, ATP gamma S, and GTP gamma S all induced two CCK binding affinity states, which in the presence of 1 mM ATP were Kd = 74 pM for high-affinity sites and Kd = 4.3 nM for low-affinity sites: AMP-PCP did not induce two affinity states. GDP at 10 microM had no effect on CCK binding but potentiated the effect of ATP. GTP gamma S, in addition to inducing high- and low-affinity states, also elicited a significant concentration-dependent reduction in the total number of measurable CCK receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Nucleoside diphosphate kinase associated with rat pancreatic membranes regulates CCK receptor affinity. 797 49

Isolated pancreatic islets from rats and humans express a plasmalogen-preferring ATP-stimulatable, Ca(2+)-independent phospholipase A2 (ASCI-PLA2) enzyme which participates in the glucose-stimulated hydrolysis of arachidonate from membrane phospholipids and in insulin secretion. Here we report that clonal insulin-secreting HIT beta-cells contain substantial amounts of endogenous plasmalogens and express a similar ASCI-PLA2 activity with the following properties: (1) Enzymatic activity as well as glucose-induced eicosanoid release and insulin secretion are inhibited by a mechanism-based suicide substrate directed towards ASCI-PLA2. (2) HIT cell ASCI-PLA2 is selectively activated and protected against thermal denaturation by ATP. (3) The magnitude of ASCI-PLA2 activation by the nonhydrolyzable ATP analog AMP-PCP is similar to that by ATP. (4) The ATP concentrations required to activate ASCI-PLA2 fall within physiologic ranges in the presence of Mg2+. (5) ADP induces a concentration-dependent attenuation of the activation of ASCI-PLA2 by ATP. HIT cell ASCI-PLA2 exhibited an apparent isoelectric point of 7.5 on chromatofocusing analysis and was quantitatively adsorbed to an ATP-agarose matrix and selectively desorbed from this column by ATP. Mono-Q anion-exchange analysis of the active ATP-agarose eluant yielded a peak of ASCI-PLA2 activity associated with a single protein band with an apparent molecular mass of 40 kDa. Similar chromatographic behavior of the rat pancreatic islet ASCI-PLA2 activity was observed during sequential ATP-agarose and Mono-Q anion-exchange steps. These results indicate that HIT cells express an ASCI-PLA2 similar to the analogous islet enzyme and suggest that expression of this enzyme and of its preferred plasmalogen substrates may be a general property of insulin-secreting beta-cells.
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PMID:Characterization of an ATP-stimulatable Ca(2+)-independent phospholipase A2 from clonal insulin-secreting HIT cells and rat pancreatic islets: a possible molecular component of the beta-cell fuel sensor. 800 9


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