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)

Cyclic nucleotide phosphodiesterase activities in soluble Neurospora crassa mycelial extracts were resolved into two peaks, phosphodiesterase I and II, by chromatography on DEAE-cellulose columns. Phosphodiesterase I hydrolysed cyclic AMP and cyclic GMP equally well. Phosphodiesterase II was active on cyclic GMP but scarcely active on cyclic AMP. Phosphodiesterase I was resolved by gel filtration and sucrose-density-gradient centrifugation into three peaks having molecular weights of about 57 000, 125 000 and 225 000. This suggests that this enzyme activity has at least three aggregation forms, tentatively defined as monomeric, dimeric and tetrameric. Similarly, phosphodiesterase II was resolved into two forms, having molecular weights of about 170 000 and 320 000. Evidence on the interconversion between phosphodiesterase I forms was obtained.
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PMID:Cyclic nucleotide phosphodiesterase activities in Neurospora crassa. 628 7

DNA polymerase has been purified approximately 2000-fold from Mycobacterium tuberculosis H37Rv. The purified preparation was homogeneous by electrophoretic criteria and has a molecular weight of 135 000. The purified enzyme resembles Escherichia coli polymerase I in its properties, being insensitive to sulfhydryl drugs and possessing 5',3'-exonuclease activity in addition to polymerase and 3',5'-exonuclease activities. However, it differs from the latter in its sensitivity to higher salt concentration and DNA intercalating agents such as 8-aminoquinoline. The polymerase exhibited maximal activity between 37--42 degrees C and pH 8.8--9.5. The polymerase was stable for several months below 0 degree C. However, the 5',3'-exonuclease activity was more labile. The effects of different metal ions, polyamines and drugs on the polymerase activity are presented.
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PMID:Purification and properties of DNA polymerase from Mycobacterium tuberculosis H37Rv. 678 93

Virions of fish lymphocystis disease virus (FLDV), a member of the iridovirus family, were isolated directly from lymphocystis disease lesions of individual flatfishes and purified by sucrose and subsequent cesium chloride gradient centrifugation to homogeneity as judged by electron microscopy. The isolated FLDV DNAs appear to be heterogeneous in size. Contour length measurements of 43 DNA molecules gave an average length of 49 +/- 23 microns, corresponding to 93 +/- 44 X 10(6) D. Molecular weight estimations of FLDV DNA by restriction enzyme analysis resulted in only 64.8 X 10(6) D indicating an excess length of the DNA of about 50%. FLDV DNA was sensitive to lambda 5'-exonuclease and to E. coli 3'-exonuclease III without preference of any one terminal DNA restriction fragment. Denaturation and reannealing experiments of FLDV DNA resulted in the formation of circular DNA molecules of 34.25 microns contour length (= 65.22 X 10(6) D). This result suggests that FLDV DNA contains directly repeated sequences at both ends and that it is terminally redundant. FLDV DNA is methylated in cytosine. FLDV DNA did not hybridize with frog virus DNA indicating that the two iridoviruses are not closely related to each other. Restriction enzyme analysis and Southern blot hybridizations revealed that FLDV isolates can be classified into two different strains: FLDV strain 1 occurs in flounders and plaice, whereas strain 2 is usually found in lesions of dabs.
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PMID:Analysis of the genome of fish lymphocystis disease virus isolated directly from epidermal tumours of pleuronectes. 685 94

A procedure of the isolation of platelets from the blood of adult sheep (Ovis aries L. var domestica) is reported. This procedure is based on differential centrifugation and a specific lysis for elimination of erythrocytes. We have obtained platelets with a purity at least 99% and a relative high yield (2.3 +/- 0.4 g wet/l whole blood identical to 235 +/- 40 mg platelet proteins/l whole blood). After disruption, homogenisation and ultracentrifugation onto a discontinuous sucrose gradient (1.6 M, 1.1 M, 1.0 M and 0.6 M sucrose), four fractions were obtained. We have separated, for the first time, a particulate preparation enriched in the whole sheep plasma membrane. This fraction was characterized by: (i) the typical membrane morphology as shown by electron micrographs; (ii) the highest activities in membrane marker enzymes such as bis(p-nitrophenyl)phosphate phosphodiesterase (EC 3.1.16.1) and 5'-dTMP-p-nitrophenyl ester phosphodiesterase (EC 3.1.3.35), and the relatively low activity for marker enzymes associated to other subcellular fractions; (iii) the highest sialic acid, cholesterol and phospholipid concentrations. The chemical composition of the platelet membrane isolated is: total proteins, 49%; lipids, 47%; carbohydrates, congruent to 3.4% (the content of hexoses is twice as high as that of hexosamines and sialic acid). The similarities and differences of this preparation with others from several sources are discussed.
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PMID:Isolation, characterization and chemical composition of the membrane from sheep platelets. 731 94

Drosophila Rrp1 includes a carboxy-terminal region homologous to Escherichia coli exonuclease III which is sufficient to repair both oxidative and alkylation damage to DNA. An apurinic/apyrimidinic endonuclease activity intrinsic to Rrp1 was characterized previously. In this work, the 3'-phosphodiesterase and 3'-phosphatase activities of Rrp1 are demonstrated and characterized. Phosphoglycolate- and phosphate-modified DNA 3'-termini are formed by oxygen radical induced DNA cleavage. To demonstrate the 3'-phosphodiesterase activity of Rrp1, a 3'-phosphoglycolate-terminated oligonucleotide substrate was generated by site-specific cleavage of a unique GpC dinucleotide by iron(II) bleomycin. Removal of the terminal phosphoglycolate is detected by mobility shift on a DNA sequencing gel. Rrp1 cleaves the phosphoglycolate and releases a product with a 3'-hydroxyl terminus. Phosphoglycolate is removed more readily than the 3'-terminal dGMP residue. Rrp1 phosphodiesterase activity is not inhibited by 120 mM NaCl, while the 3'-exonuclease is reduced 25-fold. Using a 3'-phosphate-terminated oligonucleotide, the phosphatase activity of Rrp1 is at least 25-fold lower than its phosphodiesterase or apurinic endonuclease, and 56-fold lower than exonuclease III activity on the identical substrate. Rrp1 3'-phosphatase is reduced 25-fold by 80 mM NaCl. These results were confirmed using an assay that measures the ability of Rrp1 to stimulate DNA synthesis on circular DNA substrates nicked by various DNA damage treatments. In that assay, Rrp1 poorly repairs 3'-phosphate-terminated nicks introduced by micrococcal nuclease. The significance of these enzymatic properties for the biological of Rrp1 is discussed.
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PMID:Characterization of the nuclease activity of Drosophila Rrp1 on phosphoglycolate- and phosphate-modified DNA 3'-termini. 753 50

We characterized cyclic nucleotide phosphodiesterases isolated from rat mesangial cells and assessed their roles in regulating cellular cyclic nucleotide levels. Three peaks of phosphodiesterase activity were eluted by a linear sodium acetate gradient from a Q Sepharose column loaded with the mesangial cell extract. The first peak activity was stimulated by Ca(2+)-calmodulin and inhibited by calmodulin-stimulated phosphodiesterase inhibitors but not by a selective cGMP specific phosphodiesterase V inhibitor. The second, minor activity peak was stimulated by cyclic GMP and inhibited by EHNA [erythro-9-(2-hydroxy-3-nonyl)-adenine], a selective inhibitor of cyclic GMP-stimulated phosphodiesterase II. The last peak activity was not inhibited by cyclic GMP but selectively inhibited by rolipram [4-(3-cyclopentyloxy-4-methoxyphenyl)-2-pyrrolidene] or Ro 20-1724 [4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone], inhibitors of cyclic AMP specific, cyclic GMP insensitive phosphodiesterase IV. Based on their order of chromatographic elution, kinetic properties and sensitivity to allosteric agents and inhibitors, the peak 1, 2 and 3 correspond to phosphodiesterase I, II and IV. The basal cyclic GMP level was raised more effectively by selective inhibitor of phosphodiesterase I than phosphodiesterase II. In contrast, the atrial natriuretic factor-induced cyclic GMP elevation was potentiated more effectively by selective inhibitors of phosphodiesterase II than phosphodiesterase I. The forskolin-induced cyclic AMP increase was greatly potentiated by selective phosphodiesterase IV inhibitors but not by other phosphodiesterase inhibitors. These data suggest that phosphodiesterase I and II are responsible for cyclic GMP hydrolysis whereas phosphodiesterase IV is mainly responsible for cyclic AMP hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Cyclic nucleotide phosphodiesterase isozymes in rat mesangial cells. 778 11

Drosophila Rrp1 has several tightly associated enzymatic activities, including double-strand DNA 3'-exonuclease, apurinic/apyrimidinic endonuclease, 3'-phosphatase, and 3'-phosphodiesterase. The carboxyl-terminal third of Rrp1, homologous to Escherichia coli exonuclease III, is sufficient to repair oxidative and alkylation-induced DNA damage in vivo. Using a screen for partial complementation of repair-deficient E. coli, we isolated three mutants of the nuclease domain of Rrp1: T462A, K463Q, and L484P, that protect against methyl methanesulfonate (MMS)-induced but not t-BuO2H-induced DNA damage. Thr-462 and Lys-463 are highly conserved residues found in a cluster of 5 conserved amino acids (LQETK), while Leu-484 is poorly conserved. Gln-460 Glu-461, Thr-462, and Lys-463 and Leu-484 were altered by site-directed mutagenesis using a plasmid including the entire Rrp1 gene and mutant proteins were purified. Mutants of the three residues Glu-461, Thr-462, and Lys-463 demonstrate 8-200-fold lower phosphodiesterase specific activity than wild-type Rrp1. E461A has a 30-fold reduction in AP endonuclease and is MMS-sensitive, but all other mutants have near-normal AP endonuclease and are MMS-resistant. Glu-461 appears to be essential for the nuclease function for Rrp1. Lys-463 and, to a lesser extent, Thr-462 influence the substrate specificity of the Rrp1 nuclease.
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PMID:Single amino acid changes alter the repair specificity of Drosophila Rrp1. Isolation of mutants deficient in repair of oxidative DNA damage. 779 76

Cytosine arabinoside monophosphate (araCMP) at the 3' terminus of DNA constitutes a lesion that impedes further synthesis by DNA polymerase alpha (DNA pol alpha). A biochemical assay has been designed to detect 3'-->5'-exonucleases in cell extracts that remove the 3'-araCMP lesion in an oligonucleotide template-primer and permit subsequent extension by DNA pol alpha. The major 3'-->5'-exonuclease activity in human myeloblast extracts has been purified, and gel filtration chromatography of the purified enzyme indicates that the exonuclease has an apparent native molecular mass of 52 kDa. Incubation of the enzyme with a 5'-32P-labeled araCMP template-primer results in exonucleolytic degradation of the primer exclusively in the 3'-->5' direction, demonstrating that the enzyme is a 3'-->5'-exonuclease. The products of the 3'-->5'-exonuclease reaction are 5'-mononucleotides. The apparent rate of araCMP removal by the exonuclease is approximately the same as the rate of deoxynucleoside monophosphate (dNMP) removal. Furthermore, the apparent rates of 3'-terminal excision are approximately the same whether the oligomer is hybridized to a complementary oligonucleotide, or not, indicating that the enzyme has both single- and double-stranded 3'-->5'-exonuclease activity. The enzyme does not possess 5'-->3'-exonuclease activity, nor is it associated with DNA polymerase activity. In addition, the enzyme does not cleave 3'-phosphoryl-terminated DNA, and it does not cleave RNA. The enzymatic characteristics of the isolated 3'-->5'-exonuclease indicate that it is distinct from previously identified mammalian deoxyribonucleases.
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PMID:Identification of a 3'-->5'-exonuclease that removes cytosine arabinoside monophosphate from 3' termini of DNA. 820 43

The deoxyribooligonucleotide 5'-d(CTCACATGTACACTCT) was reacted separately with the chiral diol epoxide isomers 7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha- epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-anti-BPDE)] and 7 alpha, 8 beta-dihydroxy-9 beta, 10 beta-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrene [(-)-anti-BPDE)], to produce the modified oligonucleotides 5'-d(CTCACATGBPDETACACTCT). Adducts in which either (+)-anti-BPDE or (-)-anti-BPDE are covalently bound via their C10 positions by trans addition to the exocyclic amino group of the single G residues were isolated and purified by HPLC methods. Snake venom phosphodiesterase (SVPD, phosphodiesterase I), which hydrolyzes DNA from the 3'-OH terminus to the 5'-end, digests the (+)-trans-anti-BPDE-oligonucleotide adducts at a significantly faster rate than that of the sterically different (-)-trans-anti-BPDE-oligonucleotide adducts. However, using spleen phosphodiesterase (SPD, phosphodiesterase II), which hydrolyzes DNA in the 5'-->3' direction, the opposite stereoselective resistance to digestion is observed. Using shorter BPDE-modified oligonucleotides as standards, the enzyme stall sites have been defined by gel electrophoresis methods; the most digestion-resistant phosphodiester linkage is the 5'-d(...T-G*...)-3' bond in the case of (+)-trans-BPDE-modified oligonucleotide adducts for both enzymes, SVPD and SPD (the starred G denotes the site of BPDE modification). In the case of the (-)-trans-BPDE-modified oligonucleotide adducts, the phosphodiester bond on the 3'-side of the modified G [5'-d(...G*-T...)-3'] is most resistant to digestion by both enzymes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Opposite stereoselective resistance to digestion by phosphodiesterases I and II of benzo[a]pyrene diol epoxide-modified oligonucleotide adducts. 821 49

The kinetic and functional characteristics of I and II forms of DNA-dependent DNA-polymerases of Acholeplasma laidlawii PG-8 have been studied. It is stated that I form of DNA polymerase possesses 5'-3'-exonuclease activity and is a typical replicase; II form of DNA-polymerase possesses both 5'-3'-polymerase and 3'-5'-exonuclease activity and is, evidently, a reparase. Both forms of enzyme give preference to poly(U)- and poly(A)-matrices having extremely high activity on these polymers. The enzymatic reactions realized by both forms of DNA-polymerases are described by the first-order equation. The calculated Michaelis-Menten constants equaled 180 and 250 microM for I and II forms of polymerases, respectively. It indicates that affinity to substrate in II form of polymerase is one-third higher than in I form of enzyme.
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PMID:[The kinetic and functional characteristics of DNA-dependent DNA-polymerases in Acholeplasma laidlawii PG-8]. 849 2


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