Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.4.1 (phosphodiesterase)
18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A highly purifed preparation of rat intestinal phosphodiesterase II (oligonucleate 3'-nucleotidohydrolase, EC 3.1.4.18) has been studied using a synthetic substrate, thymidine 3'(2,4-dinitrophenyl) phosphate. The enzyme was most active between pH 6.1 and pH 6.7 and was inhibited by Cu2+ and Zn2+ but unaffected by EDTA, Mg2+, Co2+, and Ni2+. The reaction rate decreased at high levels of enzyme because of competitive inhibition by deoxythymidine 3'-phosphate, a reaction product, which showed a Ki of 2-10(-5) M. The molecular weight of the enzyme by gel-filtration was 150 000-170 000. In electrofocusing experiments multiple peaks of activity were found at pH 3.4, 4.2-4.5and 7.2. Polyacrylamide gel electrophoresis of freshly purified phosphodiesterase II showed up to 10 protein bands in the gels. If the preparations were stored at 4 degrees C for some time only one or two bands appeared. Investigation of the reaction of rat intestinal phosphodiesterase II with a number of possible phosphodiesterase substrates indicated that the enzyme required a nucleoside 3'-phosphoryl residue for the initiation of hydrolysis. Thus compounds such as NAD, ATP, bis-(p-nitrophenyl)phosphate, thymidine 5'-(p-nitrophenyl)phosphate, glycerylphosphorylcholine, guanylyl-(2' leads to 5')-adenosine and 3',5'-cyclic AMP which contain phosphodiester bonds, nevertheless were not substrates for the enzyme. The enzyme was inhibited reverisbly by p-chloromercuribenzoate and p-chloromercuriphenylsulfonate and inactivated irreversibly by iodoacetic acid. Activity of the phosphodiesterase II was reduced to 50% by incubation with 2.0-10(-3)--5.0-10(-3) M iodoacetate for 20--30 min at 24 degrees C at pH 5.0--6.1. Iodoacetamide had no effect. The degree of inactivation by iodoacetate was reduced by the presence of a substrate for the enzyme or, more effectively by deoxythymidine 3'-phosphate, a competitive inhibitor. It is concluded that iodoacetic acid alkylates an essential residue at the active centre of the enzyme.
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PMID:Rat intestinal phosphodiesterase II. Properties of the highly purified enzyme and its inactivation by iodoacetic acid. 1 24

The spectroscopic properties of the 3-thioamide analogues of coenzymes NAD and NADH (sNAD and sNADH) have been investigated in order to obtain information about their conformational properties. In particular, ultraviolet absorption and circular dichroism properties of solutions in phosphate buffer pH 7 and ethanol were studied. Also equimolar mixtures of AMP and sNMN(H), obtained by cleaving the coenzymes with phosphodiesterase, were investigated using the same solvents. The appearance of a couplet around 260 nm, which is not present for the mixture of sNMN and AMP, suggests a stacking interaction of the two aromatic moieties in sNAD. This conclusion is further substantiated by a hyperchroism of the ultraviolet absorption band in the 260-nm region in both sNAD and sNADH. The comparison of the ultraviolet and circular dichroic properties of intact and cleaved coenzymes in the different solvent systems makes it possible to single out the bands which are more sensitive to conformation changes (i.e. to open-stacking equilibrium) and those which are sensitive to solvent effects only.
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PMID:Circular dichroic properties and conformation of thionicotinamide dinucleotides. 20 82

The dermal cells in grey, xanthic, and white goldfish integuments were cytochemically characterized for the following enzymatic activities: tyrosinase, DOPA-oxidase, cytochrome oxidase, monoamine oxidase, peroxidase, non-specific esterase, cholinesterase, NAD-diaphorase, NADP-diaphorase, aryl sulfatase, nucleotide phosphodiesterase, beta-glucuronidase, acid phosphatase, alkaline phosphatase, adenosine triphosphatase, thiamine pyrophosphatase, glucose-6-phosphatase, aldolase, as well as succinate, malate, isocitrate, glutamate, glucose-6-phosphate, 6-phosphogluconate, alpha-glycerophosphate, alcohol, lactate, and beta-hydroxybutyrate dehydrogenases. It was found that the epidermis was a significant barrier to the access of cytochemical reaction substrates. Removal of the epidermal barrier provided dermal cell localizations of enzymatic activities which were reproducible. Further, alterations in reaction times and temperatures from the mammalian methodology provided conditions fe various integumental cells were compared for possible interrelationships. The basic foundations for future work with the dermis of poikilothermic vertebrates on an experimental basis were established. In addition, a previously undescribed non-pigmented dermal cell, the "x"-cell, was found to have enzymatic characteristics similar to both melanophores and lipophores. The "x"-cell may be the common precursor of both types of pigment cells.
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PMID:Cytochemical characterization of goldfish (Carassius auratus L.) dermis with special reference to the pigment cells. 82 86

Gonad-stimulating substance (GSS) secreted from radial nerves induces meiotic maturation of starfish oocytes by stimulating production of 1-methyladenine (1-MeAde) in ovarian follicle cells. We have previously shown that cAMP mediates the action of GSS on 1-MeAde synthesis by starfish ovarian follicle cells. The present study examines the possible involvement of guanine nucleotide-binding regulatory proteins (G-proteins) and adenylate cyclase in the action of GSS on 1-MeAde production by starfish (Asterina pectinifera) follicle cells. GSS slightly stimulated adenylate cyclase activity in crude membrane preparations of follicle cells. GTP markedly enhanced this action of GSS in a dose-dependent manner. Nonhydrolyzable GTP analogs such as guanosine 5'-O-(3-thiotriphosphate) and 5'-guanylylimidodiphosphate, NaF, and forskolin also stimulated adenylate cyclase activity. In addition, chorela toxin (CT) stimulated adenylate cyclase activity in membrane preparations in the presence of NAD and GTP. Unlike adenylate cyclase, phosphodiesterase activity was not influenced by GSS. When crude membranes of follicle cells were incubated with [alpha-32P]NAD in the presence of CT and pertussis toxin, 45-kDa and 41-kDa proteins were ADP-ribosylated, respectively, suggesting the presence of two types (stimulatory and inhibitory) of G-proteins. It is concluded that G-proteins and adenylate cyclase play an important role in the action of GSS on 1-MeAde production by starfish ovarian follicle cells.
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PMID:Involvement of G-proteins and adenylate cyclase in the action of gonad-stimulating substance on starfish ovarian follicle cells. 184 1

When the homogenate prepared from immature rat testes was incubated with [32P]NAD, several proteins (90, 39 and 20 kDa) were ADP-ribosylated in the absence of bacterial toxins. This observation suggested the existence of an endogenous ADP-ribosyltransferase and substrates. The data that the digested product by phosphodiesterase of ADP-ribosylated 20 kDa protein was 5'-AMP suggested that 20 kDa protein was mono(ADP-ribosyl)ated. In addition, the mono(ADP-ribosyl)ation of 20 kDa protein was enhanced by guanine nucleotides such as GTP, GDP and GTP[gamma S], and decreased by the concentrations of 10 mM Mg2+. In contrast, the incorporation of ADP-ribose moiety from NAD to both 90 and 39 kDa proteins was not changed by guanine nucleotides. On the other hand, mono(ADP-ribosyl)ation of 20 kDa protein was not observed in the homogenate prepared from other tissues of the same rats. Furthermore, we found that mono(ADP-ribosyl)ation of 20 kDa protein was decreased with the maturation of the rats and that an endogenous mono(ADP-ribosyl)transferase and 20 kDa protein were located in the nuclei.
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PMID:Reduction of mono(ADP-ribosyl)ation of 20 kDa protein with maturation in rat testis: involvement of guanine nucleotides. 189 5

We used pertussis toxin to study the mechanism(s) by which divalent cations lower cellular cAMP content in bovine parathyroid cells. In cultured parathyroid cells, high extracellular Ca2+ (5 mM) or Mg2+ (5-10 mM) lowers dopamine-stimulated cAMP content by 70-90%. Pertussis toxin (0.5 microgram/ml) totally blocks the inhibitory effects of Ca2+ and Mg2+ on cAMP content. Ba2+ and Sr2+ (5 mM) also lower cAMP content by 80-90%, and this effect is, likewise, blocked by pertussis toxin. Pretreatment with pertussis toxin had no effect on the release of cAMP into the extracellular fluid. The toxin also did not modify phosphodiesterase activity in sonicates of parathyroid cells (42.68 +/- 3.26 vs. 47.00 +/- 2.82 pmol cAMP hydrolyzed/10(6) cells.20 min in control and toxin-treated cells, respectively). Moreover, addition of the phosphodiesterase inhibitor isobutyl-methylxanthine did not modify the inhibition of dopamine-stimulated cAMP accumulation by 5 mM Ca2+ in control cells (85% vs. 86% inhibition, respectively, with and without isobutylmethylxanthine). Pertussis toxin-catalyzed ADP ribosylation in homogenates of control cells demonstrated the presence of two substrates with mol wt of 40K and 41K. Preexposure of cells to pertussis toxin overnight resulted in the complete loss of both substrates on subsequent ADP ribosylation with [32P]NAD. Pertussis toxin pretreatment did not enhance adenylate cyclase activity indirectly via reducing the extracellular Ca2+-induced rise in cytosolic Ca2+, since the cytosolic Ca2+ level at 5 mM Ca2+ was about 60% higher in pertussis toxin-treated than in control cells (531 +/- 85 vs. 326 +/- 35 nM; P less than 0.05). In addition, ionomycin had no significant effect on cellular cAMP levels in control cells despite increasing the cytosolic Ca2+ concentration to levels as high as 1700 nM at 10(-5) M. Thus, changes in cytosolic Ca2+ phosphodiesterase activity, or efflux of cAMP from the cell cannot explain the inhibition of cAMP accumulation by divalent cations or the reversal of this effect by pertussis toxin. Instead, the present data suggest that extracellular divalent cations modulate the formation of cellular cAMP in parathyroid cells by a process involving a pertussis toxin-sensitive guanine nucleotide regulatory protein, presumably inhibition of adenylate cyclase by Gi via a receptor-like mechanism.
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PMID:Divalent cations suppress 3',5'-adenosine monophosphate accumulation by stimulating a pertussis toxin-sensitive guanine nucleotide-binding protein in cultured bovine parathyroid cells. 246 88

Ventricular myocytes isolated from the hypertrophied hearts of thyrotoxic adult rats have an increase in mean protein content per myocyte (6.3 +/- 0.2 vs. 4.4 +/- 0.2 ng) compared with euthyroid cells. Viability and adenine nucleotide profiles are similar in both populations, but NAD content of the hyperthyroid myocytes is depressed (4.9 +/- 0.2 vs. 5.5 +/- 0.2 nmol/mg for controls) and UTP is higher (1.2 +/- 0.09 vs. 0.9 +/- 0.04 nmol/mg). Binding of (-)-[125I]iodocyanopindolol to intact hyperthyroid myocytes is increased by 42% compared with controls, with no change in the dissociation constant (Kd). This elevation in beta-receptor number is correlated to enhanced beta-agonist-induced adenosine 3',5'-cyclic monophosphate (cAMP) production. The half-maximal effective concentration (EC50) for the euthyroid isoproterenol dose-response curve is 2.14 x 10(-7) M but is decreased to 2.51 x 10(-8) M in hyperthyroid cardiac cells. Basal adenylate cyclase activity is apparently not affected by thyroid hormones, since basal cAMP levels for both groups are identical (5 pmol/mg) and both rise roughly twofold in the presence of a phosphodiesterase inhibitor. Forskolin-induced cAMP production and cAMP-specific phosphodiesterase activity are similar as well. In contrast to beta-adrenergic response, there are no significant differences in alpha 1-antagonist [3H]prazosin binding parameters between hyperthyroid and euthyroid cardiomyocytes.
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PMID:Hyperthyroid adult rat cardiomyocytes. I. Nucleotide content, beta- and alpha-adrenoreceptors, and cAMP production. 248 Jul 17

The possible role of adenosine 3',5'-cyclic monophosphate (cAMP) in the mechanism of the acute inhibitory effects of nicotinamide and analogues on brush-border membrane (BBM) phosphate transport was investigated. Compared with basal values, cAMP content of rat renal proximal tubule suspensions was elevated two- to fivefold when incubated at 37 degrees C for 1 h with nicotinamide, 5-methylnicotinamide, or picolinamide at 1-3 mM and in the presence of a phosphodiesterase inhibitor. Thymidine had no effect on cAMP content. There was significant and specific inhibition of BBM transport of phosphate when proximal tubules were incubated with either nicotinamide or picolinamide at concentrations that increased tubule cAMP content. Thymidine had no effect on BBM transport of phosphate. These findings were independent of the dietary Pi intake of the rats. The absence of any effect of thymidine on phosphate transport strongly suggests that inhibition of poly(adenosine diphosphate ribose) polymerase does not play a role in nicotinamide action on phosphate transport. The change in phosphate transport induced by nicotinamide occurred with no change in NAD content. These findings indicate that an increase in cAMP, rather than NAD, is the important change that may mediate the acute inhibition of Na(+)-dependent phosphate transport by nicotinamide.
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PMID:Increased cAMP in proximal tubules is acute effect of nicotinamide analogues. 255 65

Binding of polyvalent antigens to IgE present in Fc epsilon receptors on the surface of mast cells and the RBL-2H3 cell line triggers the exocytotic release of allergic mediators. Preincubation of RBL-2H3 cells with cholera toxin (CT) was found to potentiate greater than or equal to 2- to 3-fold the rate and final amount of antigen-induced secretion of [3H]serotonin and N-acetyl beta-D-glucosaminidase. This was accompanied by a more variable increase in the initial rate of antigen-triggered formation of inositol phosphates. The holotoxin was required for potentiation, as neither the A nor the B subunit was effective when added separately. Four observations indicate that cAMP was not the primary effector of the augmentation of secretion caused by CT: (i) culture conditions were found in which CT caused large increases in secretion but very modest (or no) increases in cAMP; (ii) under other conditions, progressive increase in [CT] caused a maximum 2.5- to 3-fold increase in cAMP followed by a return to basal levels, whereas the secretory response saturated and remained stable; (iii) permeant cAMP analogs consistently enhanced secretion at low doses and inhibited at higher doses, but the peak enhancement was always much less than that achieved by an optimal dose of CT; (iv) the selective phosphodiesterase inhibitor Ro 20-1724 exhibited similar biphasic dose-response curves, the maximum enhancement again being small compared to that caused by CT itself. Both in vitro and in vivo, CT catalyzed transfer of ADP-ribose from NAD to two membrane proteins that comigrated on NaDodSO4/polyacrylamide gel electrophoresis with two CT substrates in other cell types, and these were identified by immunoblotting as Gs alpha. These results suggest that ADP-ribosylation of a cholera toxin substrate potentiates IgE-mediated secretion from RBL-2H3 cells by a largely cAMP-independent route.
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PMID:Cholera toxin potentiates IgE-coupled inositol phospholipid hydrolysis and mediator secretion by RBL-2H3 cells. 284 4

In order to exert its antitumor effects, the C-nucleoside tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide) is converted to the dinucleotide TAD (thiazole-4-carboxamide adenine dinucleotide), an inhibitor of IMP dehydrogenase (IMPD). With few exceptions, sensitive tumors (such as the P388 leukemia) have been found to accumulate substantially more of this inhibitory dinucleotide than resistant strains (exemplified by the colon 38 carcinoma). Previous studies have attributed this difference to a depressed capacity to synthesize TAD on the part of tumors refractory to tiazofurin. In the present study, a second contributory factor has been identified, viz. an enhanced ability to degrade preformed TAD. This degradation has been traced to a soluble phosphodiesterase present at high levels in tumors naturally resistant to tiazofurin. Using standard techniques, this TAD-phosphodiesterase has been purified 200-fold from the colon 38 carcinoma. The activity so purified readily hydrolyzed TAD and ADP-ribose, but exhibited a comparatively weak activity toward NAD and thymidine-5'-monophosphate-nitrophenyl ester. ADP-Ribose was also an excellent inhibitor of the hydrolysis of TAD. It is concluded, on the basis of these results, that TAD-phosphodiesterase plays an important role in the expression of the oncolytic activity of tiazofurin. The suggestion is also made that ADP-ribose may be the natural substrate for this enzyme.
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PMID:Studies on the mechanism of action of tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide). VI. Biochemical and pharmacological studies on the degradation of thiazole-4-carboxamide adenine dinucleotide (TAD). 287 71


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