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 novel phosphodiesterase was purified from cultured tobacco cells to a state which appeared homogeneous on polyacrylamide gel electrophoresis. The enzyme hydrolyzed various phosphodiester and pyrophosphate bonds, including p-nitrophenyl thymidine 5'-phosphate, p-nitrophenyl thymidine 3'-phosphate, cyclic nucleotides, ATP, NAD+, inorganic pyrophosphate, dinucleotides, and poly(adenosine diphosphate ribose), which is a polymer synthesized from NAD+. However, it did not hydrolyze highly polymerized polynucleotides. The molecular weight of the native enzyme was estimated as 270 000 to 280 000 by gel filtration on Sephadex G-200 and Bio-Gel A-5m. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the enzyme was composed of subunits with molecular weights calculated to be 75 000. The enzyme did not require divalent cations for activity being fully active in the presence of ethylenediaminetetraacetic acid. The pH optimum for the enzyme was approximately 6 with p-ni-trophenyl thymidine 5'-phosphate or adenosine cyclic 3',5'monophosphate, and 5.3 with NAD+. Double reciprocal plots of the initial velocity against the concentration of p-nitrophenyl thymidine 5'-phosphate gave two apparent Km values of 0.17 and 1.3 mM, suggesting the presence of at least two active sites.
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PMID:A novel phosphodiesterase from cultured tobacco cells. 0 41

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

A simple method, involving NAD+-Sepharose chromatography, was developed for the preparation of snake venom phosphodiesterase (EC 3.1.4.1) almost free from 5'-Nucleotidase (EC 3.1.3.5). Using an NAD+-Sepharose 4B column, phosphodiesterase was eluted in the unadsorbed fraction, whereas 5'nucleotidase was strongly adsorbed. The latter enzyme was desorbed when 0.2 M sodium bicarbonate buffer containing 1mM beta-NADH was used as a solvent. The affinity column could be used at least four times without any decrease of potency, and the method was applicable for the preparation of phosphodiesterase from the venoms of rattlesnake (Crotalus adamanteus) and Japanese mamushi (Agkistrodan halys blomhoffi).
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PMID:A simple method for preparation of snake venom phosphodiesterase almost free from 5'-nucleotidase. 16 89

4-Nitrophenyl and 2-napthyl monoesters of phenylphosphonic acid have been synthesized, and an enzyme catalyzing their hydrolysis was resolved from alkaline phosphatase of a commerical calf intestinal alkaline phosphatase preparation by extensive ion-exchange chromatography, chromatography on L-phenylalanyl-Sepharose with a decreasing gradient of (NH4) 2SO4, and gel filtration. Detergent-solubilized enzyme from fresh bovine intestine was purified after (NH4)2SO4 fractionation by the same technique. The purified enzyme is homogeneous by polyacrylamide gel electrophoresis and sedimentation equilibrium centrifugation. It has a molecular weight of 108,000, contains approximately 21% carbohydrate, and has an amino acid composition considerably different from that reported from alkaline phosphatase from the same tissue. The homogeneous intestinal enzyme, an efficient catalyst of phosphonate ester hydoolysis but not of phosphate monoester hydrolysis, was identified as a 5'-nucleotide phosphodiesterase by its ability to hydrolyze 4-nitrophenyl esters of 5'-TMP but not of 3'-TMP. Also consistent with this identification was the ability of the enzyme to hydrolyze 5'-ATP to 5'-AMP and PPi, NAD+ to 5'-AMP and NMN, TpT to 5'-TMP and thymidine, pApApApA to 5'-AMP, and only the single-stranded portion of tRNA from the 3'-OH end. Snake venom 5'-nucleotide phosphodiesterase also hydrolyzes phosphonate esters, but 3'-nucleotide phosphodiesterase of spleen and cyclic 3',5'-AMP phosphodiesterase do not. Thus, types of phosphodiesterases can be conveniently distinguished by their ability to hydrolyze phosphonate esters. As substrates for 5'-nucleotide phosphodiesterases, phosphonate esters are preferable to the more conventional esters of nucleotides and bis(4-nitrophenyl) phosphate because of their superior stability and ease of synthesis. Furthermore, the rate of hydrolysis of phosphonate esters under saturating conditions is greater than that of the conventional substrates. At substrate concentrations of 1 mM the rates of hydrolysis of phosphonate esters and of nucleotide esters are comparable and both superior to that of bis(4-nitrophenyl) phosphate.
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PMID:Hydrolysis of phosphonate esters catalyzed by 5'-nucleotide phosphodiesterase. 17 Sep 64

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

1. The effects of changes in the cytoplasmic [NADH]/[NAD+] ratio on the efficacy of glucagon to alter rates of metabolism in isolated rat hepatocytes were examined. 2. Under reduced conditions (with 10mM-lactate), 10nM-glucagon stimulated both gluconeogenesis and urea synthesis in isolated hepatocytes from 48h-starved rats; under oxidized conditions (with 10mM-pyruvate), 10nM-glucagon had no effect on either of these rates. 3. The ability of glucagon to alter the concentration of 3':5'-cyclic AMP and the rates of glucose output, glycogen breakdown and glycolysis in cells from fed rats were each affected by a change in the extracellular [lactate]/[pyruvate] ratio; minimal effects of glucagon occurred at low [lactate]/[pyruvate] ratios. 4. Dose-response curves for glucagon-mediated changes in cyclic AMP concentration and glucose output indicated that under oxidized conditions the ability of glucagon to alter each parameter was decreased without affecting the concentration of hormone at which half-maximal effects occurred. 5. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (0.05 mM) significantly reversed the inhibitory effects of pyruvate on glucagon-stimulated glucose output. 6. For exogenously added cyclic [3H]AMP(0.1 mM), oxidized conditions decreased the stimulatory effect on glucose output as well as the intracellular concentration of cyclic AMP attained, but did not alter the amount of cyclic [3H]AMP taken up. 7. The effects of lactate, pyruvate, NAD+ and NADH on cyclic AMP phosphodiesterase activities of rat hepatocytes were examined. 8. NADH (0.01--1 MM) inhibited the low-Km enzyme, particularly that which was associated with the plasma membrane. 9. The inhibition of membrane-bound cyclic AMP phosphodiesterase by NADH was specific, reversible and resulted in a decrease in the maximal velocity of the enzyme. 10. It is proposed that regulation of the membrane-bound low-Km cyclic AMP phosphodiesterase by nicotinamide nucleotides provides the molecular basis for the effect of redox state on the hormonal control of hepatocyte metabolism by glucagon.
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PMID:Responsiveness to glucagon by isolated rat hepatocytes controlled by the redox state of the cytosolic nicotinamide--adenine dinucleotide couple acting on adenosine 3':5'-cyclic monophosphate phosphodiesterase. 21 54

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

Poly(ADP-ribose) built from NAD+ on histones and other nuclear proteins by poly(ADP-ribose) polymerase is involved in repair, replication, gene expression, recombination, and chromatin remodeling in embryogenesis. Such nuclear processes are believed to be facilitated by opening up of condensed chromatin structures and by removal of histones from DNA at damaged sites as well as at origins of replication and transcription initiation sites. In addition, poly(ADP-ribosyl)ation might be involved in the up or down regulation of the activity of key nuclear enzymes. Poly(ADP-ribose) is rapidly synthesized at sites containing DNA strand breaks and is then rapidly degraded (half-life 0.5-5 min) by poly(ADP-ribose)glycohydrolase. High-resolution polyacrylamide gel electrophoresis is used in this study to analyze the rate of consumption of [32P]NAD+, the rate of formation of poly(ADP-ribose) molecules, and the rate of appearance of ADP-ribose, AMP, and phosphoribosyl-AMP, the catabolites of poly(ADP-ribose) in isolated nuclei from mouse cells in culture. Our method permits direct loading of aliquots of nuclei at time intervals on the polyacrylamide gel. The action of poly(ADP-ribose) glycohydrolase that degrades the polymer starts at less than 2 min from polymer formation. A poly(ADP-ribose) phosphodiesterase present in mammalian cell nuclei begins degrading poly(ADP-ribose) or unincorporated NAD+ and free ADP-ribose at 10 min. Mammalian phosphodiesterase is identified as an enzyme more important than previously thought which might degrade poly(ADP-ribosyl)ated proteins but also recycle the ADP-ribose produced from di- to poly(ADP-ribosyl)ated proteins by glycohydrolase into utilizable AMP units.
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PMID:Poly(ADP-ribose) synthesis and degradation in mammalian nuclei. 132 75

A phosphodiesterase from bull seminal plasma was purified to homogeneity. The purification procedure involved sequential column chromatographies on DEAE-Sephadex A-50, ConA-Agarose, chromatofocusing and AMP-Agarose. The final yield was about 20% with a 3000-fold purification. As indicated by chromatofocusing, the enzyme is an acidic protein (pI approximately 4.6) and owing to its interaction with Concanavalin A it is also a glycoprotein. The SDS-PAGE showed that the purified phosphodiesterase seemed to be constituted of a single polypeptide chain of about 125 kDa. The enzyme did not show an absolute substrate specificity. Thus, it was able to hydrolyze 4-nitrophenyl ester of 5'-TMP (but not of 3'-TMP), cAMP, nucleic acids as well as NAD+, ADP and ATP. According to its enzymatic properties, bull seminal plasma phosphodiesterase is to be considered an oligonucleate 5'-nucleotidohydrolase. In addition the seminal plasma phosphodiesterase also showed phosphonate esterase activity.
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PMID:Bull seminal plasma phosphodiesterase. Purification and general properties. 133 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.
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PMID:Mono ADP-ribosylation of transducin catalyzed by rod outer segment extract. 151 16


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