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)

Labeling with [3H]galactose was employed to isolate a glycosylphosphatidylinositol from rat hepatocytes which might be involved in the action of insulin. The polar head group of this glycosylphosphatidylinositol was generated by phosphodiesterase hydrolysis with a phosphatidylinositol-specific phospholipase C from Bacillus cereus. By Dowex AG1 x 8 chromatography the polar head group could be separated into three radioactive peaks eluting at 100 mM (peak I), 200 mM (peak II) and 500 mM (peak III) ammonium formate, respectively. Peak III was the most active as an inhibitor of the cAMP-dependent protein kinase. Treatment of peak III with alkaline phosphatase markedly reduced its activity on cAMP-dependent protein kinase. When peaks I, II or III were treated with alkaline phosphatase and analyzed again by Dowex AG1 x 8 chromatography, the radioactivity eluted with the aqueous fraction. The above results indicate that the polar head group of the insulin-sensitive glycosylphosphatidylinositol from rat hepatocytes exists in three different phosphorylated forms and that the biological activity of this molecule depends on its phosphorylation state.
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PMID:Different phosphorylated forms of an insulin-sensitive glycosylphosphatidylinositol from rat hepatocytes. 304 67

Calmodulin (CaM), a multifunctional calcium binding protein with no known enzymatic activity, has been purified to homogeneity from bovine adrenal cortex. The purification included anion exchange on DE-52 cellulose, ammonium sulfate precipitation, and separation by molecular sieving on Sephadex G-150. The yield of CaM from 900 g of whole adrenal was 150 mg. Adrenocortical CaM showed a molecular weight of 18,000 on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, an isoelectric point of 4.1, and demonstrated a characteristic shift in mobility on polyacrylamide gels in the presence of calcium. The spectral properties of adrenocortical CaM differed slightly from those of CaM isolated from bovine brain. Minor differences were observed in peptide maps and amino acid composition between adrenocortical and brain CaM, but adrenocortical CaM contained a single trimethyl-lysine residue characteristic of all mammalian forms of CaM isolated to date. Adrenocortical CaM is biologically active in the stimulation of activator-deficient phosphodiesterase, and showed a half-maximal effective concentration (EC50) of 3 nM for stimulation of adenylate cyclase from Bordetella pertussis.
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PMID:Purification and properties of calmodulin from adrenal cortex. 397 May 28

1. The enzyme fraction obtained from the pH5 enzyme of non-lactating bovine mammary gland between 40 and 100% ammonium sulphate saturation markedly inhibited the AMP-incorporating activity of rat liver nucleotide-incorporating enzyme. This inhibitory effect has been attributed to high nuclease activity which can be partially removed by adsorption of the enzyme fraction on to calcium phosphate gel. 2. The degradation action of the calcium phosphate-purified enzyme is confined mainly to the terminal trinucleotide sequence -pCpCpA of tRNA, its effect being analogous to that of venom phosphodiesterase. This enzyme is heat labile and very readily loses its degradative activity. 3. Treatment of the enzyme fraction with Macaloid results in complete removal of the phosphodiesterase, leaving an enzyme capable of incorporating AMP into tRNA. 4. Transfer RNA extracted from non-lactating bovine mammary gland in the presence of polyvinyl sulphate and Macaloid is able to accept amino acids with an efficiency 30% of that shown by lactating bovine mammary-gland tRNA isolated under identical conditions.
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PMID:Competing addition and hydrolysis of the cytidylylcytidylyladenosine terminal residues of transfer ribonucleic acid isolated from the non-lactating bovine mammary gland. 432 Dec 70

1. The purification of a nuclease from rat-liver mitochondria is described. The mitochondria are rendered soluble by treatment with Triton X-100 and, after fractionation with ammonium sulphate and acetone, the active fraction is further purified by chromatography on DEAE-cellulose and Sephadex G-75 to give a purification of over 700-fold. 2. The purified enzyme was only very slightly contaminated with deoxyribonuclease II, phosphodiesterase and phosphomonoesterase. The individual activities of these enzymes did not exceed 0.1% of the activity of the liver nuclease. 3. The purified enzyme attacked RNA more rapidly than denatured DNA and hydrolysed native DNA more slowly than denatured DNA. 4. There is some evidence to suggest that the nucleolytic activity of the purified preparation towards native DNA, denatured DNA and RNA is associated with a single protein. 5. The enzyme is relatively labile but is stabilized in the presence of 20% (w/v) glycerol or 10mm-2-mercaptoethanol.
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PMID:The purification from rat liver of a nuclease hydrolysing ribonucleic acid and deoxyribonucleic acid. 591 28

The characteristics of soluble and membrane-bound glutamine synthetase (GS) from Rhodospirillum rubrum were compared with those of the enzyme located in situ (measured in detergent-treated cells). The results suggest that in vivo GS may be associated with, or bound to, the chromatophore membranes. GS was found to reversibly associate and dissociate from purified chromatophores as a function of the ionic strength of the buffer or the Mg2+ concentration. Solubilized GS was purified to homogeneity and found to be similar to the GS of enteric bacteria in that its molecular weight was about 600,000 and it had one type of subunit of 51,000 molecular weight. Removal of GS from the membrane had no effect on the Km values for the substrates of the biosynthetic reaction, but it did have a substantial effect on both its Mg2+ requirement (the Km increased 10-fold) and the sensitivity of the gamma-glutamyl transferase reaction to the inhibitor methionine sulfoximine (the I0.5 decreased from 1,500 to 60 microM). Both observations suggest that the active site of GS is influenced by its association with the membrane. GS activity was shown to respond to NH4+, phosphodiesterase, Mg2+, and adenylylation cofactors in a manner identical to that of the GS of the coliform bacteria, suggesting that the former may also respond to adenylylation and deadenylylation. Finally, R. rubrum GS was also inhibited by NH4+ by a newly observed, as yet undefined, system.
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PMID:Evidence for a glutamine synthetase-chromatophore association in the phototroph Rhodospirillum rubrum: purification, properties, and regulation of the enzyme. 613 14

Mycobacterium avium was previously shown to be dependent upon ammonia or glutamine as a nitrogen source. In an effort to assess the physiology of ammonia assimilation by M. avium, a characterization of its glutamine synthetase was performed. The enzyme from M. avium was purified by streptomycin sulfate treatment, ammonium sulfate precipitation, and affinity chromatography. The enzyme was unusual in that it had a pH optimum of 6.4 and maximum enzyme activity was obtained between 50 and 60 degrees C as shown by the transferase assay. The glutamine synthetase activity from batch-cultured cells decreased with increasing concentration of ammonium chloride in the range of 0.25-5 mumol/mL of medium, which demonstrated a response to environmental supply of a nitrogen source. The mycobacterial enzyme was similar to the other bacterial glutamine synthetases in terms of molecular weight and sedimentation coefficient which were 600 000 and 19.5 S, respectively, and enzyme activity was lost by treatment with a glutamate analog, methionine sulfoximine. The isoelectric point was, however, pH 4.5. Treatment of the enzyme with snake venom phosphodiesterase resulted in an increase in specific activity. AMP was released by the phosphodiesterase treatment, thus demonstrating that M. avium glutamine synthetase was regulated by adenylylation modification.
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PMID:Glutamine synthetase from Mycobacterium avium. 614 81

Some properties of the soluble phosphatidylinositol phosphodiesterase (monophosphatidylinositol inositolphosphohydrolase, EC 3.1.4.10) of rabbit iris smooth muscle are described. Studies on its subcellular distribution showed that in this tissue the phosphodiesterase is not exclusively cytosolic. Thus, under our experimental conditions about 58% of the enzyme activity was found in the soluble fraction and the remainder was particulate. When the latter was treated with deoxycholate about 59% of the enzyme activity, compared to 86% of that of ATPase, was still bound to the particulate fraction. The kinetic properties of the enzyme (30--50% (NH4)2SO4 fraction) were examined. Maximum breakdown was 7.7 mumol/h per mg protein and occurred at pH 5.6. The products of [14C]arachidonic acid-labelled phosphatidylinositol were 1,2-diacylglycerol and a mixture of 86% myoinositol 1-phosphate and 14% myoinositol 1,2-(cyclic)phosphate. The enzyme has an absolute requirement for Ca2+. Addition of Ba2+, La3+, Mg2+, Mn2+, EGTA or EDTA at 0.05--5 mM concentrations; Sr2+ at higher concentrations (greater than 0.25 mM) markedly inhibited the phosphodiesterase activity and this inhibition was completely reversed by Ca2+. The enzyme is specific for the phosphoinositides.
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PMID:Studies on the properties of a soluble phosphatidylinositol-phosphodiesterase of rabbit iris smooth muscle. 625 Jun 28

Calmodulin was purified to apparent homogeneity from sea urchin spermatozoa by heat-treatment at 85 degrees C, ammonium sulphate precipitation at pH 4.2, DEAE-Sephacel chromatography and gel filtration on Sephadex G-100. Approximately 8.3 micrograms calmodulin were recovered per 10(10) sperm cells. The sperm calmodulin had an apparent molecular weight of 17 800. The purified calmodulin activated calmodulin-deficient phosphodiesterase from pig coronary arteries, with half-maximal activation occurring at approximately 40 ng calmodulin/ml. Trifluoperazine also inhibited the sperm calmodulin activity. These results demonstrate that calmodulin is present in high amounts in sea urchin spermatozoa, and that it is essentially the same as the calmodulin isolated from various other tissues.
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PMID:Purification and characterization of calmodulin from sea urchin spermatozoa. 625 34

1. A rat brain supernatant and microsomal fraction contained a phospholipase A1 enzyme which hydrolysed phosphatidylinositol at pH 8 in the absence of calcium. Triolein and phosphatidylcholine were not attacked under the same incubation conditions. 2. No evidence could be obtained for a phospholipase A2 in the microsomal preparation, and in the presence of Ca2+ the release of fatty acid observed was due to phosphatidylinositol phosphodiesterase followed by diacylglycerol lipase action. 3. Brain phosphatidylinositol phosphodiesterase showed extensive activity in the alkaline range (7-8.5) as well as at pH 5-5.5. The activity at higher pH values required higher calcium concentrations and disappeared on purification of the soluble enzyme by ammonium sulphate fractionation. 4. In general the ratio between inositol 1,2-(cyclic)phosphate and inositol 1-phosphate produced by phosphodiesterase action decreased with increasing pH.
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PMID:The catabolism of phosphatidylinisitol by an EDTA-insensitive phospholipase A1 and calcium-dependent phosphatidylinositol phosphodiesterase in rat brain. 627 69

Extracts of Chlorella pyrenoidosa, Euglena gracilis var. bacillaris, spinach, barley, Dictyostelium discoideum and Escherichia coli form an unknown compound enzymically from adenosine 5'-phosphosulphate in the presence of ammonia. This unknown compound shares the following properties with adenosine 5'-phosphoramidate: molar proportions of constituent parts (1 adenine:1 ribose:1 phosphate:1 ammonia released at low pH), co-electrophoresis in all buffers tested including borate, formation of AMP at low pH through release of ammonia, mass and i.r. spectra and conversion into 5'-AMP by phosphodiesterase. This unknown compound therefore appears to be identical with adenosine 5'-phosphoramidate. The enzyme that catalyses the formation of adenosine 5'-phosphoramidate from ammonia and adenosine 5'-phosphosulphate was purified 1800-fold (to homogeneity) from Chlorella by using (NH(4))(2)SO(4) precipitation and DEAE-cellulose, Sephadex and Reactive Blue 2-agarose chromatography. The purified enzyme shows one band of protein, coincident with activity, at a position corresponding to 60000-65000 molecular weight, on polyacrylamide-gel electrophoresis, and yields three subunits on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of 26000, 21000 and 17000 molecular weight, consistent with a molecular weight of 64000 for the native enzyme. Isoelectrofocusing yields one band of pI4.2. The pH optimum of the enzyme-catalysed reaction is 8.8. ATP, ADP or adenosine 3'-phosphate 5'-phosphosulphate will not replace adenosine 5'-phosphosulphate, and the apparent K(m) for the last-mentioned compound is 0.82mm. The apparent K(m) for ammonia (assuming NH(3) to be the active species) is about 10mm. A large variety of primary, secondary and tertiary amines or amides will not replace ammonia. One mol.prop. of adenosine 5'-phosphosulphate reacts with 1 mol.prop. of ammonia to yield 1 mol.prop. each of adenosine 5'-phosphoramidate and sulphate; no AMP is found. The highly purified enzyme does not catalyse any of the known reactions of adenosine 5'-phosphosulphate, including those catalysed by ATP sulphurylase, adenosine 5'-phosphosulphate kinase, adenosine 5'-phosphosulphate sulphotransferase or ADP sulphurylase. Adenosine 5'-phosphoramidate is found in old samples of the ammonium salt of adenosine 5'-phosphosulphate and can be formed non-enzymically if adenosine 5'-phosphosulphate and ammonia are boiled. In the non-enzymic reaction both adenosine 5'-phosphoramidate and AMP are formed. Thus the enzyme forms adenosine 5'-phosphoramidate by selectively speeding up an already favoured reaction.
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PMID:Purification and properties of adenylyl sulphate:ammonia adenylyltransferase from Chlorella catalysing the formation of adenosine 5' -phosphoramidate from adenosine 5' -phosphosulphate and ammonia. 627 7


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