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Query: HUMANGGP:030741 (calmodulin-dependent phosphodiesterase)
 89 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Trypsin-treated Ca2+/calmodulin-dependent phosphodiesterase (CA2+-PDE), which had lost its sensitivity to Ca2+-calmodulin, was inhibited by various calmodulin antagonists, trifluoperazine, chlorpromazine, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) and aminoalkyl chain analogues of W-7 (A-3, A-4, A-5, I-240, A-6, A-7). These inhibitory effects were less than those on calmodulin-activated Ca2+-PDE. The ability of these compounds to inhibit trypsin-treated Ca2+-PDE correlated well with the inhibitory effect on calmodulin-activated Ca2+-PDE. W-7 inhibited trypsin-treated Ca2+-PDE in a competitive fashion with respect to cyclic GMP and the Ki value was 300 microM. The inhibition of trypsin-treated Ca2+-PDE by W-7 (300 microM) or A-7 (100 microM) was overcome by the addition of excess calmodulin. Trypsin-treated Ca2+-PDE can bind to W-7-coupled cyanogen bromide-activated Sepharose 4B in the presence of 1 mM EGTA. These results suggest that Ca2+-PDE possesses a binding site for calmodulin antagonists and that the binding site for these antagonists on this enzyme may be structurally similar to the binding site on calmodulin itself.
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PMID:Direct interaction of calmodulin antagonists with Ca2+/calmodulin-dependent cyclic nucleotide phosphodiesterase. 609 52

In many cell systems, the permeability of membrane junctions is modulated by the cytoplasmic level of free Ca++. To examine whether the calcium-dependent regulatory protein calmodulin is involved in this process, the ability of anticalmodulin drugs to influence the cell-to-cell passage of injected current and an organic tracer was tested using standard intracellular glass microelectrode techniques. Several antipsychotics and local anesthetics were found to block junctional communication in the epidermis of the beetle Tenebrio molitor. Treatment of the epidermis with chlorpromazine (0.25 mM) raised intercellular resistance two- to threefold within 20 to 25 min; cell-to-cell passage of electrical current was abolished within 41 +/- 5 min. Loss of electrotonic coupling was accompanied by a block in the cell-to-cell movement of the organic tracer carboxyfluorescein. The reaction is fully reversible, with normal electrotonic coupling being restored within 2 to 4 hr. Other antipsychotics and local anesthetics had similar effects on cell coupling. The order of potency found was: trifluoperazine greater than thioridazine greater than D-butaclamol greater than chlorprothixine = chlorpromazine greater than L-butaclamol greater than dibucaine greater than tetracaine. The relative uncoupling potencies of these drugs correlate well with their known ability to inhibit calmodulin-dependent phosphodiesterase activity. Other anesthetic compounds, procaine and pentobarbital, did not block cell-to-cell communication. Altering the extracellular Ca++ concentration did not affect the rate of uncoupling by antipsychotics, while chelation of extracellular Ca++ with EGTA raised electrotonic coupling. The effect of three metabolic inhibitors on coupling was also examined. Iodoacetate uncoupled the epidermal cells while DNP and cyanide did not. These results are discussed in terms of possible mechanisms by which calmodulin may control junctional communication in this tissue.
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PMID:Drugs that block calmoduLin activity inhibit cell-to-cell coupling in the epidermis of Tenebrio molitor. 612 25

Ca2+-regulated guanylate cyclase in ciliary membranes from Paramecium contained tightly bound calmodulin. Antisera against calmodulin from Tetrahymena and soybean inhibited enzyme activity. EGTA did not easily release calmodulin; however, La3+ inhibited guanylate cyclase by dissociation of calmodulin. While La could not replace Ca in the activation of guanylate cyclase, it substituted for Ca2+ in the activation of calmodulin-dependent phosphodiesterase from pig brain independently of whether homologous or Paramecium calmodulin was used. After removal of endogenous calmodulin from guanylate cyclase, reconstitution was achieved with calmodulin from Paramecium, Tetrahymena, pig brain, and soybean. Ca2+-binding proteins lacking trimethyllysine like calmodulin from Dictyostelium, parvalbumin, and troponin C failed to restore enzyme activity. The properties of the native and reconstituted guanylate cyclase/calmodulin complex were compared. Reassociation of calmodulin with its target enzyme was weak since all calmodulin remained in the supernatant after a single centrifugation. While most enzyme characteristics remained unchanged in the reconstituted complex, the inhibition by Ca greater than 100 microM was of a mixed-type compared to noncompetitive inhibition in the native enzyme. The regulation of the enzyme by cations was also altered. Whereas Ca was the most potent and specific activator of the native enzyme, in the reconstituted system Sr was far more effective.
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PMID:Calcium/calmodulin-regulated guanylate cyclase of the excitable ciliary membrane from Paramecium. Dissociation of calmodulin by La3+: calmodulin specificity and properties of the reconstituted guanylate cyclase. 613 52

3T3-L1 adipocytes contain both soluble and particulate cAMP phosphodiesterases which can be distinguished by several criteria. Particulate phosphodiesterase activity of 3T3-L1 adipocytes, but not undifferentiated fibroblasts, was selectively increased by incubation of cells with insulin or lipolytic hormones. Particulate cAMP phosphodiesterase activity from 3T3-L1 adipocytes was very sensitive to inhibition by cilostamide in an apparently competitive fashion. Particulate activity from undifferentiated 3T3-L1 fibroblasts or supernatant activity from either type of cell was much less sensitive to cilostamide. On the other hand, supernatant cAMP phosphodiesterase activity from both undifferentiated fibroblasts and 3T3-L1 adipocytes was very sensitive to inhibition by Ro-20-1724 in an apparently competitive fashion. Ro-20-1724 was not an effective inhibitor of particulate activity from either type of cell. In fractions from 3T3-L1 adipocytes, isobutylmethylxanthine (IBMX) effectively inhibited both supernatant and particulate cAMP phosphodiesterase activities. In addition, however, IBMX was relatively more specific in inhibiting supernatant calmodulin-activated cGMP phosphodiesterase activity than supernatant calmodulin-independent or particulate cGMP phosphodiesterase activities. In intact 3T3-L1 adipocytes, cilostamide enhanced lipolysis in the absence or presence of isoproterenol and had no effect on cAMP content in the presence of low concentrations of isoproterenol. Ro-20-1724 increased lipolysis to a lesser extent than cilostamide and did not enhance isoproterenol-stimulated lipolysis, but did increase isoproterenol-stimulated accumulation of cAMP to a greater extent than cilostamide. Like cilostamide, Ro-20-1724 did not enhance accumulation of cAMP in the absence of isoproterenol. IBMX enhanced lipolysis and cAMP accumulation with or without isoproterenol. Taken together, these results support the idea that although particulate and soluble low Km phosphodiesterases influence cAMP content, the particulate enzyme may be more important in the metabolism of cAMP involved in the regulation of lipolysis. Since combinations of Ro-20-1724 and cilostamide were not as effective as IBMX in increasing cAMP content, perhaps the calmodulin-dependent phosphodiesterase, which is selectively inhibited by IBMX, is also involved in the regulation of total cell cAMP content.
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PMID:Selective effects of phosphodiesterase inhibitors on different phosphodiesterases, adenosine 3',5'-monophosphate metabolism, and lipolysis in 3T3-L1 adipocytes. 620 9

Calmodulin-dependent cyclic nucleotide phosphodiesterase was purified from bovine brain to apparent homogeneity by a new procedure involving DEAE-cellulose, Affi-Gel blue, calmodulin-Sepharose 4B, and Sephadex G-200 column chromatographies. The enzyme was purified more than 3,000-fold from the brain extracts with greater than 12% yield. The purified phosphodiesterase could be activated 10- to 15-fold by calmodulin and Ca2+ to a specific enzyme activity of more than 300 mumol of cAMP hydrolyzed/min/mg of protein. Molecular weight of the enzyme was determined to be 115,800 by the sedimentation equilibirum method or 124,000 from the sedimentation constant and Stokes radius of the protein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the enzyme showed a single protein band with an apparent molecular weight of 58,000. These results suggested that the calmodulin-dependent phosphodiesterase from bovine brain has a subunit structure of alpha2. Molecular weight of the complex of calmodulin and phosphodiesterase was the complex of calmodulin and phosphodiesterase was also calculated from the sedimentation constant and Stokes radius to be 159,000. Since calmodulin has a molecular weight of about 17,000, the result indicated that the stoichiometry of the complex is calmodulin2 alpha2. The catalytic subunit of cylic AMP-dependent protein kinase was found to catalyze the phosphorylation of the purified phosphodiesterase with the incorporation of 2 mol of phosphate/mol of the enzyme.
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PMID:Purification and properties of bovine brain calmodulin-dependent cyclic nucleotide phosphodiesterase. 624 53

Divalent metals used to support phosphodiesterase (EC 3.1.4.-) activity have been found to influence the substrate and enzyme specificity of many phosphodiesterase inhibitors in studies of the hydrolysis of cyclic AMP and cyclic GMP by the calmodulin-dependent and cyclic AMP-specific phosphodiesterases from bovine heart. Many compounds displayed marked differences in substrate specificity and inhibitory potency in the presence of Mg2+, as compared with Mn2+, when studied with the unactivated form of calmodulin-dependent phosphodiesterase, while few compounds displayed differences in the presence of calmodulin. With a single divalent metal, marked differences in inhibitory potency and substrate specificity were also observed in the absence or presence of calmodulin suggesting that alterations in calmodulin and/or Ca2+ levels may greatly affect the response to phosphodiesterase inhibitors. Divalent metals did not alter the effects of inhibitors on the hydrolysis of cyclic AMP by the cyclic AMP-specific phosphodiesterase, however divalent metals would probably indirectly influence the relative cellular level of cyclic AMP hydrolyzed by this enzyme, and therefore the effects of inhibitors, through metal effects on the calmodulin-dependent phosphodiesterase. No correlation was found between the inhibitory activity of the compounds, many of which were cyclic nucleotide analogs, and their ability to activate cyclic AMP-dependent or cyclic GMP-dependent protein kinases or to affect cyclic AMP-dependent protein kinase activity by displacing bound cyclic AMP.
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PMID:Effects of divalent metals on the specificity of inhibitors of the cyclic nucleotide phosphodiesterases from bovine heart. 626 Jan 97

Interactions between phosphorylase kinase (ATP:phosphorylase-b phosphotransferase, EC 2.7.1.38) and calmodulin were studied with pure preparations of muscle phosphorylase kinase, and with crude extracts from muscles of control (C57 Black) and deficient (ICR/IAn) mice, which lack muscle phosphorylase kinase activity. Calmodulin was determined by its ability to stimulate a calmodulin-dependent phosphodiesterase. The amount of calmodulin bound to phosphorylase kinase in muscle extract was estimated to a maximum of 30% of the total amount of calmodulin. In the muscle of the deficient strain a decrease of 35% in the total amount of calmodulin was observed. This correlates with the absence of the calmodulin fraction specifically bound to phosphorylase kinase. From sucrose gradient studies we demonstrated that in the presence of Ca2+ the amount of calmodulin bound to phosphorylase kinase was enhanced, compared to the control in the presence of EGTA. This observation was made both in crude extracts and in pure phosphorylase kinase preparations. Sucrose gradient also showed that muscle phosphorylase kinase can be dissociated to low molecular species when extracts are made in the presence of Ca+; this dissociation was found to be related to a Ca2+-dependent proteolytic effect.
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PMID:Calmodulin ligands. The interaction of muscle phosphorylase kinase with phosphodiesterase. Comparison of calmodulin ligands in muscle extracts from normal and phosphorylase kinase-deficient mice. 626 Feb 1

The apparent target sizes of the basal and calmodulin-dependent activities of calmodulin-activated phosphodiesterase from bovine brain were estimated using target theory analysis of data from radiation inactivation experiments. Whether crude or highly purified samples were irradiated, the following results were obtained. Low doses of radiation caused a 10 to 15% increase in basal activity, which, with further irradiation, decayed with an apparent target size of approximately 60,000 daltons. Calmodulin-dependent activity decayed with an apparent target size of approximately 105,000 daltons. The percentage stimulation of enzyme activity by calmodulin decreased markedly as a function of radiation dosage. These observations are consistent with results predicted by computer-assisted modeling based on the assumptions that: 1) the calmodulin-activated phosphodiesterase exists as a mixture of monomers which are fully active in the absence of calmodulin and dimers which are inactive in the absence of calmodulin; 2) in the presence of calmodulin, a dimer exhibits activity equal to that of two monomers; 3) on radiations destruction of a dimer, an active monomer is generated. This monomer-dimer hypothesis provides a plausible explanation for and definition of basal and calmodulin-dependent phosphodiesterase activity.
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PMID:Calmodulin-activated cyclic nucleotide phosphodiesterase from brain. Relationship of subunit structure to activity assessed by radiation inactivation. 627 Jan 50

1. Plasma membranes from ascites hepatoma cells (AH-7974, AH-130) contained much smaller amounts of calmodulin (about half) and cyclic AMP phosphodiesterase (about one-third) compared to plasma membranes of rat livers. 2. Some of calmodulin molecules in liver plasma membranes were released by repeated washing. The 'washed' liver plasma membranes showed the presence of specific binding sites for externally added calmodulin molecules (bovine brain) (N = 140 pmol/mg protein, Kd = 7.9 . 10(-8) M). The calmodulin content of AH-7974 plasma membranes was not reduced by repeated washing. The binding of calmodulin to the 'washed' AH-7974 plasma membranes was only of nonspecific nature with negative cooperativity. 3. Plasma membranes (liver and AH-7974) appeared to contain both calmodulin-dependent and calmodulin-independent phosphodiesterase, but the stimulation by externally added Ca2+ plus calmodulin was rather small. Externally added calmodulin-dependent phosphodiesterase (bovine brain) was bound more to 'washed' liver plasma membranes than to 'washed' AH-7974 plasma membranes. Newly bound phosphodiesterase appeared to be more sensitive to the stimulation by Ca2+ plus calmodulin in 'washed' hepatoma plasma membranes than in 'washed' liver plasma membranes. 4. Preincubation of 'washed' plasma membranes (liver and hepatoma) with calmodulin did not affect the binding of phosphodiesterase, but the sensitivity of phosphodiesterase to the stimulation by Ca2+ plus calmodulin in hepatoma plasma membranes was lost.
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PMID:Dynamics of calmodulin and cyclic AMP phosphodiesterase in plasma membranes of rat livers and ascites hepatomas. 627 Dec 50

Cyclic nucleotide phosphodiesterase activities (3',5'-cyclic nucleotide 5'-nucleotidohydrolase, EC 3.1.4.17) were found in the 40,000 X g supernatant fraction of homogenates of Xenopus laevis oocytes. In the supernatant, the ratio of the specific activity of cyclic AMP phosphodiesterase to that of cyclic GMP phosphodiesterase was 1.1 at the 1 micro substrate level. Two phosphodiesterase forms were isolated by centrifugation on sucrose gradient: a 3-4 S form hydrolyzing specificity cyclic AMP and a 6-7 S form hydrolyzing both cyclic nucleotides (cyclic AMP and cyclic GMP). The activity of the 6-7 S phosphodiesterase was characterized by its activation by 0.1 micro M calmodulin purified from beef pancreas in the presence of 50 micro M CA2+. The calmodulin dependence of this form was completely abolished in the presence of 1 mM ethyleneglycobis(beta-aminoethyl ether)-N-N,N',N'-tetraacetic acid (EGTA). Trifluoperazine at 0.1 mM inhibited both the freshly prepared crude enzyme and the partially purified 6-7 S form. On the other hand, no effect of cyclic GMP at 3 micro M was observed on cyclic AMP hydrolysis in the case of the supernatant or that of the partially purified phosphodiesterases. These data show the presence of a calmodulin-dependent phosphodiesterase in the soluble fraction of X. laevis oocytes.
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PMID:Characterization of the soluble cyclic nucleotide phosphodiesterases in Xenopus laevis oocytes. Evidence for a calmodulin-dependent enzyme. 628 Jul 70


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