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

---

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

R 79595 (N-cyclohexyl-N-methyl-2-[[[phenyl (1,2,3,5-tetrahydro-2 oxoimidazo [2,1-b]-quinazolin-7-yl) methylene] amin] oxy] acetamide) and its isomers represent a novel class of compounds with phosphodiesterase (PDE) inhibitory and cardiotonic (positive inotropic) actions. The cardiac effects of this class of compounds were investigated in the hexobarbital-depressed heart-lung preparation of the guinea-pig. After induction of heart failure (reduction of cardiac output to 25% of the initial value) cumulative addition of R 79595 or its isomers R 80122 (E-isomer) and R 80123 (Z-isomer) concentration-dependently reversed the cardiac depressant effects of hexobarbitone-Na. With regard to reconstitution of contractility and cardiac function R 80122 (E-isomer) was 10 fold more potent than R 79595 (1:1 mixture of the isomers) and nearly 100 fold more potent than R 80123 (Z-isomer). Furthermore, the cardiotonic action of the most potent isomer (R 80122) was compared to the effects of several positive inotropic reference compounds. The order of cardiotonic potency was as follows: (-)-adrenaline > R 80122 = adibendan > digitoxin > milrinone = enoximone > theophylline. Adibendan (EC50 value: 6.7 +/- 1.8 x 10.-8 mol/l), which showed cardiotonic effects in the same concentration range as R 80122 (EC50 value: 6.1 +/- 1.3 x 10(-8) mol/l), was significantly (p < 0.01) less effective than R 80122 with respect to the maximally induceable increase in cardiac output (CO). The cardiotonic effects of R 80122 could be observed in the low concentration range of 3 x 10(-8) to 1 x 10(-6) mol/l, whereas enoximone (EC50 value: 1.2 +/- 0.1 x 10(-5) mol/l) and milrinone (EC50 value: 8.9 +/- 3.5 x 10(-6) mol/l) elicited positive inotropic effects at 100 fold higher concentrations. Digitoxin was 10 fold less and theophylline was 300 fold less potent than R 80122 with regard to reconstitution of heart function. The cardiotonic effects of R 80122 were not accompanied by an increase in heart rate as found with milrinone, theophylline or (-)-adrenaline in this model. Furthermore, the PDE inhibitory effect of R 79595 and its E-isomer R 80122 were investigated in partially purified isoenzymes from guinea-pig ventricles. The IC50 values of R 79595 and R 80122 on PDE I-IV were compared to the IC50 values of adibendan, milrinone, enoximone and theophylline. The selectivity of an inhibitor for PDE III was evaluated by division of its IC50 values on PDE I, II and IV by the IC50 value on PDE III. R 80122 was the most potent and selective PDE III inhibitor.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Cardiac effects of R 79595 and its isomers (R 80122 and R 80123) in an acute heart failure model. A new class of cardiotonic agents with highly selective phosphodiesterase III inhibitory properties. 147 Feb 28

Transducin is the retinal rod outer segment (ROS)-specific G protein coupling the photoexcited rhodopsin to cyclic GMP-phosphodiesterase. The alpha subunit of transducin is known to be ADP-ribosylated by bacterial toxins. We investigated the possibility that transducin is modified in vitro by an endogenous ADP-ribosyltransferase activity. By using either ROS, cytosolic extract of ROS or purified transducin in the presence of [alpha-32P]nicotinamide adenine dinucleotide (NAD+), the alpha and beta subunits of transducin were found to be radiolabeled. The labeling was decreased by snake venom phosphodiesterase I (PDE I). The modification was shown to be mono ADP-ribosylation by analyses on thin layer chromatography of the PDE I-hydrolyzed products which revealed only 5'AMP residues. In addition we report that sodium nitroprusside activates the ADP-ribosylation of transducin.
...
PMID:Mono ADP-ribosylation of transducin catalyzed by rod outer segment extract. 151 16

The DNA synthesizing subunit (alpha-subunit) of DNA polymerase-alpha from calf thymus was separated from the other three subunits by immunoaffinity chromatography. The enzymatic properties of the alpha-subunit were characterized and compared with those of the four-subunit complex. Free alpha-subunit behaved in many respects like the four-subunit polymerase-primase. It was inhibited by aphidicolin and butylanilino-deoxyATP and catalyzed DNA synthesis on both gapped duplex DNA as well as primed single-stranded DNA with a preference of gapped DNA. The alpha-subunit is a quasi-processive enzyme with a processivity for about 9 nucleotides incorporated per single primer binding event. This is 2-fold lower than the processivity of the four-subunit complex. Despite this moderate processivity, free alpha-subunit was able to synthesize long stretches of DNA on singly primed natural psi X174am16 DNA. The accuracy of DNA synthesis of the free alpha-subunit was determined by using the psi X174am16 reversion assay to be 1 error per 50,000 nucleotides incorporated. An in vitro accuracy of 1 error in 54,000 nucleotides incorporated was measured in parallel for the four-subunit complex. Thus, the smaller subunits do not contribute to the overall accuracy of DNA polymerase-alpha. Consistent with this result is the observation that the polymerase to 3'----5'-exonuclease ratio was less than 1 to 2,500,000. Therefore, there is no evidence for the action of a cryptic proofreading activity with the alpha-subunit of DNA polymerase-alpha of mammalian origin.
...
PMID:The DNA synthesizing subunit of polymerase-primase from calf thymus. 153 34

DNA replication occurs in vivo with very high processivity, meaning that the replication complex assembles at the origin(s) of replication and then performs template-directed synthesis of DNA over virtually the entire genome without dissociation. Such processivity also characterizes reconstituted replication holoenzyme complexes in vitro. However, the isolated DNA polymerases are much less processive, especially under physiological conditions. In this paper we monitor the degree of processivity displayed by the bacteriophage T4-coded DNA polymerase while in its proofreading mode by asking whether an isolated polymerase can "edit-out" the 3'-terminal nucleotide from the primer (using the 3'----5'-exonuclease activity of the polymerase) and then switch into the synthesis mode without dissociating from the DNA template. This "switch experiment" is accomplished by using mismatched primer/template substrates as an experimental tool to mimic the situation that T4 DNA polymerase encounters after a misincorporation event has occurred. By performing experiments under single-turnover conditions (obtained using a heparin trap), we demonstrate that T4 DNA polymerase, upon encountering a misincorporated base, neither synthesizes the next base nor dissociates into solution. Instead, with a greater than 80% probability, it removes the misincorporated base and then continues synthesis in a fully processive manner. We also show that the removal of a doubly mispaired sequence from the 3'-terminus of the primer, followed by synthesis, is comparably processive. In contrast, the apparent processivity of removing a triply mispaired terminus is much reduced. Taken together, these observations are consistent with the notion that the "editing active site" of the T4 enzyme optimally accommodates only two unpaired nucleotide residues. Our results do not support the idea that the exonuclease activity of T4 DNA polymerase is highly selective for mismatched termini; they suggest instead that the dwell time at a misincorporated base determines overall editing efficiency. The integrated results of this study provide additional insight into the structure of the T4 DNA polymerase, as well as into the interactions between the polymerase and the polymerase accessory proteins that are required to provide the holoenzyme complex with full processivity.
...
PMID:Processive proofreading is intrinsic to T4 DNA polymerase. 162 15

The protein responsible for both nucleotide pyrophosphatase (EC 3.6.1.9) and alkaline phosphodiesterase I (EC 3.1.4.1) activities was purified from MOPC 315 plasmacytoma cells. A single SDS/PAGE-purified 115-kDa protein band was used to produce a rabbit polyclonal antiserum. This antibody preparation precipitated alkaline phosphodiesterase I activity, indicating that the SDS/PAGE-purified protein was nucleotide pyrophosphatase/alkaline phosphodiesterase I. When used for Western blot analysis, the antiserum detected a 115-kDa protein as well as a 220-kDa protein band. Multiple overlapping cDNA clones were isolated from a cDNA expression library screened with this anti-nucleotide pyrophosphatase/alkaline phosphodiesterase I antiserum. Sequence analysis indicated that the isolated cDNA clones encoded PC-1, a murine plasma cell differentiation antigen. To confirm the suspected enzymatic identity of PC-1, a recombinant PC-1 fusion protein was expressed in bacteria, purified, and used to produce another rabbit polyclonal antiserum. This antiserum likewise immunoprecipitated alkaline phosphodiesterase I activity and recognized the 115-kDa and 220-kDa proteins in Western blot analyses of cell extracts. Furthermore, expression of nucleotide pyrophosphatase/alkaline phosphodiesterase I corresponded directly with mRNA and protein levels of PC-1 in cells known to express different levels of nucleotide pyrophosphatase/alkaline phosphodiesterase I activity. Finally, steroid induction of enzymatic activity was mirrored by levels of PC-1 mRNA and protein expression. Together, these data indicate that the plasma cell differentiation antigen PC-1 is a membrane-bound enzyme, nucleotide pyrophosphatase/alkaline phosphodiesterase I.
...
PMID:Identification of nucleotide pyrophosphatase/alkaline phosphodiesterase I activity associated with the mouse plasma cell differentiation antigen PC-1. 164 27

The pyridazinone derivative zardaverine has recently been introduced as a potent bronchodilator in vivo and in vitro. In addition, zardaverine exerts a positive inotropic action on heart muscle in vitro. The actions of zardaverine are thought to be mediated via inhibition of phosphodiesterase (PDE) activity. Recent data suggest that there are multiple forms of phosphodiesterases and at least five different isozyme families are now recognized. In the present study, the effects of zardaverine on the different PDE isozymes were investigated in several tissues. PDE isozymes were separated by chromatography on Q-sepharose. Zardaverine inhibited the cyclic GMP-inhibitable PDE III from human platelets and the rolipram-inhibitable PDE IV from canine trachea and human polymorphonuclear (PMN) cells with IC50-values of 0.58, 0.79 and 0.17 microM, respectively. The pyridazinone derivative affected the calmodulin-stimulated PDE I, the cyclic GMP-stimulated PDE II and the cyclic GMP-specific PDE V only marginally at concentrations up to 100 microM. Zardaverine inhibits the ADP-induced aggregation of human platelets with an IC50 of 1.6 microM. This inhibition was synergistically increased by activators of adenylate cyclase such as PGE1 and forskolin. In human PMN cells, zardaverine inhibited the zymosan-induced superoxide anion generation with an IC50 of 0.40 microM. Again, this effect was increased by activators of adenylate cyclase. These data clearly demonstrate that zardaverine is a selective inhibitor of PDE III and PDE IV isozymes.
...
PMID:Zardaverine as a selective inhibitor of phosphodiesterase isozymes. 164 20

Modified deoxynucleosides 2'-deoxy-beta-L-uridine, beta-L-thymidine, alpha-L-thymidine, 2'-deoxy-beta-L-adenosine and 2'-deoxy-alpha-L-adenosine were synthesized and assembled as homooligomers, respectively: octa-beta-L-deoxyuridylates, octa beta-L and alpha-L-thymidylates and tetra beta-L and alpha-L-deoxyadenylates. These unnatural oligomers were then substituted with an acridine derivative. The binding studies of these modified oligonucleotides with D-ribo- and D-deoxyribopolynucleotides were carried out by absorption spectroscopy. While beta-L-d(Up)8m5Acr, beta-L-(Tp)8m5Acr, alpha-L-(Tp)8m5Acr did not interact with poly(rA) and poly(dA), beta-L-d(Ap)4m5Acr and alpha-L-d(Ap)4m5Acr did form double and triple helices with poly(rU) and poly(dT), respectively. Their stability towards nuclease digestion was studied through comparison with that of octa-beta-D-thymidylate and tetra beta-D-deoxyadenylate covalently linked to an acridine derivative. One endonuclease (nuclease P1 from Penicillium citrinum) and two exonucleases (a 3'-exonuclease from Crotalus durissus venom and a 5'-exonuclease extracted from calf thymus) were employed. beta-L- and alpha-L-oligomers demonstrate a high resistance toward nuclease digestion.
...
PMID:Synthesis and physicochemical properties of oligonucleotides built with either alpha-L or beta-L nucleotides units and covalently linked to an acridine derivative. 165 74

The bacteriophage PRD1 DNA polymerase gene (gene I) has been cloned into the expression vector pPLH101 under the control of the lambda pL promoter. Tailoring of an efficient ribosome binding site in front of the gene by polymerase chain reaction led to a high level heat-inducible expression of the corresponding gene product (P1) in Escherichia coli cells. Expression was confirmed in vivo by complementation of phage PRD1 DNA polymerase gene mutants and in vitro by formation of the genome terminal protein P8-dGMP replication initiation complex. Expressed PRD1 DNA polymerase was purified to apparent homogeneity in an active form. DNA polymerase, 3'-5'-exonuclease, and P8-dGMP replication initiation complex formation activities cosedimented in glycerol gradient with a protein of 65 kDa, the size expected for PRD1 DNA polymerase. The DNA polymerase was active on DNase I-activated calf thymus DNA, poly(dA).oligo(dT) and poly(dA-dT) primer/templates as well as on native phage PRD1 genome. The 3'-5'-exonuclease activity was specific for single-stranded DNA and released mononucleotides. No 5'-3'-exonuclease activity was detected. The inhibitor/activator spectrum of the PRD1 DNA polymerase was also studied. An in vitro replication system with purified components for bacteriophage PRD1 was established. Formation of the P8-dGMP replication initiation complex was a prerequisite for phage DNA replication, which proceeded from the initiation complex and yielded genome length replication products.
...
PMID:Overexpression, purification, and characterization of Escherichia coli bacteriophage PRD1 DNA polymerase. In vitro synthesis of full-length PRD1 DNA with purified proteins. 165 59

We have mapped the position of the alpha-globin gene cluster in the 20- to 300-kilobase fragments of chromosomal DNA isolated from growing chicken HD3 erythroblastoid cells exposed to 4'-demethylepipodophyllotoxinthenylidene beta-D-glucoside. This epipodophyllotoxin traps functioning topoisomerase II molecules, the denaturation of which cleaves DNA and reveals their reaction sites. The DNA fragments, prepared by centrifugation in sucrose gradients, bind selectively to glass-fiber filters and are protected from lambda 5'-exonuclease, properties compatible with the presence of a topoisomerase II subunit bound to their 5' ends. Restriction enzyme cleavage of the fragments and hybridization with cloned alpha-globin-region probes reveal additional distinctive bands not seen in control DNA, allowing the localization of fragment ends near this gene cluster. The terminal regions of fragments from sucrose gradients or from field-inversion electrophoresis gels were also used to probe cloned regions of the gene cluster. Both approaches show that this cluster of three genes, which is not expressed in these cells, is located at a specific position in a approximately 20-kilobase DNA fragment. The upstream end of this fragment lies in a region that contains a site of DNA attachment to the nuclear matrix mapped by both in vivo and in vitro methods, and its downstream end is flanked by approximately 80% A + T sequences characteristic of matrix-attachment regions. These observations suggest that the DNA fragments are formed because topoisomerase II molecules can specifically and readily integrate into DNA at matrix-attachment regions and that the fragments represent entire DNA loops or domains.
...
PMID:Precise localization of the alpha-globin gene cluster within one of the 20- to 300-kilobase DNA fragments released by cleavage of chicken chromosomal DNA at topoisomerase II sites in vivo: evidence that the fragments are DNA loops or domains. 165 47

X-ray studies of the proofreading 3',5'-exonuclease site of the large (Klenow) fragment of DNA polymerase I have detected a binuclear metal complex consisting of a pentacoordinate metal (site A) which shares a ligand, Asp-355, with an octahedral metal (site B) [Freemont, P. S., Friedman, J. M., Beese, L. S., Sanderson, M. R., & Steitz, T. A. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 8924-8928; Beese, L. S., & Steitz, T. A. (1991) EMBO J. 10, 25-33]. Kinetic studies of the activation of the 3',5'-exonuclease reaction by Co2+, Mn2+, or Mg2+, at low concentrations of DNA, reveal sigmoidal activation curves for the three metal ions with Hill coefficients of 2.3-2.4 and K0.5 values of 16.6 microM, 4.2 microM, and 343 microM, respectively. The binding of Co2+ to the enzyme results in the appearance of an intense visible absorption spectrum of the metal ion with maxima at 633, 570, and 524 nm and extinction coefficients of 190, 194, and 150 M-1 cm-1, respectively, suggesting the formation of a pentacoordinate Co2+ complex. Optical titration with Co2+ yields a sigmoidal titration curve which is best fit by assuming the cooperative binding of three Co2+ ions with a K0.5 of 39.9 microM, comparable to the value of 16.6 microM obtained kinetically. Displacement of Co2+ by 1 equiv of Zn2+, which binds tightly to the A site of the 3',5'-exonuclease, shifts the optical spectrum to 524 nm and lowers the extinction coefficient to 30 -1 cm-1, indicative of octahedral coordination.2+ the formation of the binuclear complex.
...
PMID:Role of divalent cations in the 3',5'-exonuclease reaction of DNA polymerase I. 165 60


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---