Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.1 (phosphodiesterase)
 18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The PDE4A (type IV) cAMP-specific, rolipram-inhibited phosphodiesterase RPDE-6 (RNPDE4A5), when transiently expressed in COS7 cells, could be complexed with the v-Src-SH3 domain expressed as a glutathione S-transferase (GST) fusion protein. RPDE-6 did not interact with GST itself. This complex was not disrupted by treatment with high NaCl concentration together with Triton X-100. Interaction was apparently determined by the N-terminal splice region of RPDE-6, as the PDE4A splice variant RPDE-39, which differs from RPDE-6 at the extreme N-terminus, failed to associate with v-Src-SH3; met26RD1 (where RD1 is rat 'dunc-like' PDE), which has the N-terminal splice region deleted, failed to associate with v-Src-SH3, and the association of RPDE-6 and v-Src-SH3 was blocked by a fusion protein formed from the N-terminal splice region. RDPE-6 showed binding to GST fusion proteins of both the intact Src kinase and an SH2-SH3 construct but did not bind to the Src-SH2 domain or to the adaptor protein Grb-2. RPDE-6 could be co-immunoprecipitated from cytosol extracts of transfected cells by using anti-Src antiserum. RPDE-6 exhibited selectivity in binding to the SH3 domains of c-Abl, Crk, Csk, Lck, Lyn, Fyn and v-Src, with binding to the SH3 regions of the Src-related tyrosyl kinases Lyn and Fyn being the most effective. The binding of RPDE-6 to the SH3 domains of Crk, Csk and Lck led to a marked reduction in PDE activity, but no change was apparent in complexes with other species. Endogenous RPDE-6 from brain, but not endogenous RPDE-39 from testis, bound to the Src-SH3 domain. We suggest that the PDE4A splice variant RPDE-6 has a propensity for interaction with selective SH3 domains, in particular those from Src and the Src-related tyrosyl kinases Lyn and Fyn. This interaction seems to be governed by alternative splicing of the PDE4A gene, because RPDE-39, a splice variant that lacks the proline-rich N-terminal splice region of RPDE-6, does not interact with these SH3 domains. It is proposed that the binding site on RPDE-6 for SH3 domains lies within the unique first 102 residues of its N-terminal splice domain, where two motifs representing Class I SH3 binding sites with selectivity for Src kinase SH3 domains can be identified and one motif for a putative Class II SH3 binding site.
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PMID:The SH3 domain of Src tyrosyl protein kinase interacts with the N-terminal splice region of the PDE4A cAMP-specific phosphodiesterase RPDE-6 (RNPDE4A5). 876 80

A potential role for cAMP in regulating the differentiation of myoblasts has led us to examine the components of the cAMP signaling system, including the type IV, cAMP-specific phosphodiesterases. The full coding sequence of the phosphodiesterase PDE4D1 was inserted in the bacterial expression vector pGEX-KG. N- and C-terminal truncations were also placed in the same vector, allowing the expression and purification of glutathione S-transferase (GST)-PDE fusion proteins using glutathione-Sepharose. The purified PDE was active [V(max) = 318 +/- 18 nmol min(-1)(mg of protein)(-1)] and inhibited by RO 20-1724, rolipram, and MIX (IC50 values of 2, 0.4, and 40 microM, respectively). The requirement of PDE4D1 for a divalent cation was also examined. It was able to use Mg2+, Co2+, and Mn2+, but not Zn2+, suggesting that it is not a zinc hydrolase as has been proposed for other PDE types. Deletion of both C- and N-terminal regions affected the apparent native size of the enzyme. The C-terminal region was involved in dimer formation, whereas an N-terminal region was responsible for larger aggregates. Removal of the last 35 amino acids of an N-terminal 80-residue highly conserved region (UCR2) resulted in a 6-fold increase in PDE activity, providing evidence that this part of the molecule acts as an intramolecular inhibitor. The availability of a highly purified, enzymatically active protein in substantial quantities has allowed us to directly examine PDE4D1 for the first time.
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PMID:Recombinant expression of a type IV, cAMP-specific phosphodiesterase: characterization and structure-function studies of deletion mutants. 906 27

Arrestin facilitates phototransduction inactivation through binding to photoactivated and phosphorylated rhodopsin (RP). However, the specific portions of arrestin that bind to RP are not known. In this study, two different approaches were used to determine the regions of arrestin that bind to rhodopsin: panning of phage-displayed arrestin fragments against RP and cGMP phosphodiesterase (PDE) activity inhibition using synthetic arrestin peptides spanning the entire arrestin protein. Phage display indicated the predominant region of binding was contained within amino acids 90-140. A portion of this region (residues 95-140) expressed as a fusion protein with glutathione S-transferase is capable of binding to rhodopsin regardless of the activation or phosphorylation state of the receptor. Within this region, the synthetic peptide of residues 109-130 was shown to completely inhibit the binding of arrestin to rhodopsin with an IC50 of 1.1 mM. The relatively high IC50 of this competition suggests that this portion of the molecule may be only one of several regions of binding between arrestin and RP. A survey of synthetic arrestin peptides in the PDE assay indicated that the two most effective inhibitors of PDE activity were peptides of residues 111-130 and 101-120. These results indicate that at least one of the principal regions of binding between arrestin and RP is contained within the region of residues 109-130.
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PMID:Identification of regions of arrestin that bind to rhodopsin. 1005 46

For the identification of yeast genes specifying biochemical activities, a genomic strategy that is rapid, sensitive, and widely applicable was developed with an array of 6144 individual yeast strains, each containing a different yeast open reading frame (ORF) fused to glutathione S-transferase (GST). For the identification of ORF-associated activities, strains were grown in defined pools, and GST-ORFs were purified. Then, pools were assayed for activities, and active pools were deconvoluted to identify the source strains. Three previously unknown ORF-associated activities were identified with this strategy: a cyclic phosphodiesterase that acts on adenosine diphosphate-ribose 1"-2" cyclic phosphate (Appr>p), an Appr-1"-p-processing activity, and a cytochrome c methyltransferase.
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PMID:A biochemical genomics approach for identifying genes by the activity of their products. 1055 52

Bovine PDEdelta was originally copurified with rod cGMP phosphodiesterase (PDE) and shown to interact with prenylated, carboxymethylated C-terminal Cys residues. Other studies showed that PDEdelta can interact with several small GTPases including Rab13, Ras, Rap, and Rho6, all of which are prenylated, as well as the N-terminal portion of retinitis pigmentosa GTPase regulator and Arl2/Arl3, which are not prenylated. We show by immunocytochemistry with a PDEdelta-specific antibody that PDEdelta is present in rods and cones. We find by yeast two-hybrid screening with a PDEdelta bait that it can interact with farnesylated rhodopsin kinase (GRK1) and that prenylation is essential for this interaction. In vitro binding assays indicate that both recombinant farnesylated GRK1 and geranylgeranylated GRK7 co-precipitate with a glutathione S-transferase-PDEdelta fusion protein. Using fluorescence resonance energy transfer techniques exploiting the intrinsic tryptophan fluorescence of PDEdelta and dansylated prenyl cysteines as fluorescent ligands, we show that PDEdelta specifically binds geranylgeranyl and farnesyl moieties with a Kd of 19.06 and 0.70 microm, respectively. Our experiments establish that PDEdelta functions as a prenyl-binding protein interacting with multiple prenylated proteins.
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PMID:Photoreceptor cGMP phosphodiesterase delta subunit (PDEdelta) functions as a prenyl-binding protein. 1456 60

Esterase and transferase activities were analyzed simultaneously after cytosol proteins in the bovine retina were separated by microscale non-denaturing two-dimensional electrophoresis (2-DE). Esterase activity was specifically inhibited by an esterase inhibitor, 9-amino-1,2,3,4-tetra hydroacridine (tacrine), and transferase activity was specifically inhibited by a glutathione S-transferase (GST) inhibitor, 2-phenyl-1,2-benziso selenazol-3(2H)-one (ebselen). Both esterase and transferase were precipitated when ammonium sulfate was added to the cytosol up to 50% saturation (50% AS fraction), and were detected in the 50% AS fraction by using the 2-DE. After the cytosol proteins in the 50% AS fraction were separated by using non-denaturing 2-DE, polypeptides of the separated proteins were identified by peptide mass fingerprinting and post-source decay analysis by using MALDI-MS, or by immunoreactivity by using a specific antibody. The spots of esterase and transferase activities in the 2-DE pattern were identified as phosphodiesterase and GST, respectively. This simultaneous analysis of enzyme activities can be applied to screen-specific or non-specific medicines which affect enzyme activities.
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PMID:Simultaneous analysis of esterase and transferase activities in cytosol proteins from the bovine retina by using microscale non-denaturing two-dimensional electrophoresis. 1472 4

We assessed the influence of the prophylactic use of a combination of the IV beta-adrenergic blocker, esmolol, and the phosphodiesterase III inhibitor, enoximone, on postbypass hemodynamic status, inflammation, and endothelial and organ function in a prospective, randomized, placebo-controlled study in 42 patients aged >65 yr undergoing aortocoronary bypass grafting. In 21 patients, esmolol (aim: heart rate <70 bpm) plus enoximone (initial bolus of 0.5 mg/kg followed by a continuous infusion of 2.5 microg x kg(-1) x min(-1)) was started after induction of anesthesia and continued until the morning of the first postoperative day; another 21 patients received saline solution as placebo. Hemodynamics, splanchnic perfusion (gastric-arterial CO(2) gap), liver function (glutathione transferase-alpha plasma levels), renal function (creatinine clearance, urine concentrations of N-acetyl-beta-D-glucosaminidase), myocardial ischemia (creatine-kinase MB and troponin T plasma levels), inflammation (elastase, interleukin-6 and -8 plasma levels), and endothelial integrity (adhesion molecules plasma levels) were assessed at baseline, before and after cardiopulmonary bypass (CPB), and in the intensive care unit until the first postoperative day. Catecholamine requirements were significantly less in the treated than in the nontreated patients. Heart rate was significantly slower, cardiac index was higher, and gastric-arterial CO(2) gap was significantly lower in the treatment group. Troponin T, beta-N-acetyl-beta-D-glucosaminidase, glutathione transferase-alpha, and soluble adhesion molecules increased significantly in the untreated control, but remained almost normal in the esmolol+enoximone patients. Inflammatory responses (elastase/interleukins) were attenuated by esmolol+enoximone. We conclude that, in comparison to an untreated control, the prophylactic use of a combination of esmolol and enoximone in elderly patients undergoing cardiac surgery with cardiopulmonary bypass resulted in overall beneficial effects on postbypass hemodynamic status, organ function, inflammatory response, and endothelial integrity.
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PMID:The prophylactic use of the beta-blocker esmolol in combination with phosphodiesterase III inhibitor enoximone in elderly cardiac surgery patients. 2145 Oct 80

A novel gene, DD3-3, from Dictyostelium discoideum has been isolated by an mRNA differential display between a wild-type strain AX2 and a mutant HG794 which is defective in O-glycosylation. Functional analysis of the novel gene, DD3-3, was conducted by preparing a knockout mutant, DD3-3KO, and a GST:DD3-3 fusion protein. The mutant DD3-3KO cells were allowed to develop about 1.5 h earlier than the wild-type strain AX2 cells. Northern blotting analysis of the knockout mutant cells showed a remarkable downregulation of Reg A, cAMP-dependent phosphodiesterase, and overexpression of protein tyrosine kinase (PTK) during early development and its shutdown during late development. The relationship between O-glycosylation and phosphorylation involving Reg A gene is discussed.
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PMID:Functional analysis of a novel gene, DD3-3, from Dictyostelium discoideum. 1588 3

Eight furoquinoline alkaloids were purified from two plants belonging to the Rutaceae family. Kokusaginine, skimmianine, evolitrine, and confusameline were purified from Melicope confusa, and haplopine, robustine, dictamine, and gamma-fagarine from Dictamnus albus. In this study, the eight furoquinoline alkaloids were examined for inhibitory potency against human phosphodiesterase 5 (hPDE5A) in vitro. DNA encoding the catalytic domain of human PDE5A was amplified from the mRNA of T24 cells by RT-PCR and was fused to GST in an expression vector. GST-tagged PDE5A was then purified by glutathione affinity chromatography and used in inhibition assays. Of the eight alkaloids, gamma-fagarine was the most potent inhibitor of PDE5A, and its single methoxy group at the C-8 position was shown to be critical for inhibitory activity. These results clearly illustrate the relationship between PDE5A inhibition and the methoxy group position in furoquinoline alkaloids.
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PMID:Inhibitory effects of furoquinoline alkaloids from Melicope confusa and Dictamnus albus against human phosphodiesterase 5 (hPDE5A) in vitro. 1604 76

The prune gene of Drosophila melanogaster is predicted to encode a phosphodiesterase. Null alleles of prune are viable but cause an eye-color phenotype. The abnormal wing discs gene encodes a nucleoside diphosphate kinase. Killer of prune is a missense mutation in the abnormal wing discs gene. Although it has no phenotype by itself even when homozygous, Killer of prune when heterozygous causes lethality in the absence of prune gene function. A screen for suppressors of transgenic Killer of prune led to the recovery of three mutations, all of which are in the same gene. As heterozygotes these mutations are dominant suppressors of the prune-Killer of prune lethal interaction; as homozygotes these mutations cause early larval lethality and the absence of imaginal discs. These alleles are loss-of-function mutations in CG10065, a gene that is predicted to encode a protein with several zinc finger domains and glutathione S-transferase activity.
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PMID:Loss-of-function mutations in a glutathione S-transferase suppress the prune-Killer of prune lethal interaction. 1614 20


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