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)

A modification of Aurbach & Houston's enzymic method for measuring cAMP is presented. The procedure is relatively simple and in several respects new. Urinary cAMP is separated from other nucleotides and phosphate by ZnSO4-Ba(OH)2 precipitation and column chromatography. The eluate is concentrated by evaporation. Recovery at this stage is 60-82%. The cAMP from urine and the standards are dissolved in a reaction mixture and converted to 5-AMP with cyclic 3',5'-nucleotide phosphodiesterase (PDE) and further to ATP with adenylate kinase and pyruvate kinase. The ATP formed is labelled with 32P by an exchange reaction catalysed by glyceraldehyde phosphate dehydrogenase and phosphoglycerate kinase. The remaining 32P used to count the [32P]ATP in the aqueous phase. Daily human urinary cAMP excretion is 3380 +/- 836 nmol (S.D.). After an injection of 100 USP units of parathormone intravenously into a patient with idiopathic hypoparathyroidism, urinary cAMP excretion increased 40-fold above the basal concentration within 30 min. Drinking of coffee or water did not affect cAMP excretion. The limit of detection of the method is 170 pmol of cAMP, and the variation coefficient for urine ranges from 7 to 10%. When the enzymic cAMP method was compared with a radioimmunological procedure, the correlation coefficient was found to be 0.98.
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PMID:Determination of cyclic adenosine 3',5'-monophosphate in urine. 17 27

The location of 2',3'-cyclic nucleotide 2',3'-phosphodiesterase in human erythrocyte membranes was determined. This was accomplished by comparing the enzyme's accessibility with that of glyceraldehyde-3-phosphate dehydrogenase (cytoplasmic surface marker) and acetylcholinesterase (external marker) in sealed and unsealed ghosts and normal and inverted membrane vesicles. The results showed that 2',3'-cyclic nucleotide 3'-phosphodiesterase, like glyceraldehyde-3-phosphate dehydrogenase, meets several criteria for an inner (cytoplasmic) membrane location: (1) the enzyme was accessible to substrate in unsealed ghosts and inside-out vesicles but not in sealed or right-side-out vesicles, (2) latent activity in sealed ghosts could be exposed with detergent (Triton X-100), (3) activity in unsealed ghosts was gradually sequestered during resealing and could be re-exposed with detergent, and (4) the enzyme was susceptible to trypsin proteolysis only in unsealed ghosts. These results demonstrate that the active site of 2',3'-cyclic nucleotide 3'-phosphodiesterase faces the cytoplasm of erythrocytes and that the enzyme may not span the lipid bilayer of the membrane. The localization of the phosphodiesterase on the inner membrane surface of erythrocytes suggests that the similar enzyme of myelin may be embedded within the major dense line of the compact lamellae.
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PMID:Localization of 2',3'-cyclic nucleotide 3'-phosphodiesterase in human erythrocyte membranes. 627 4

The specific activity of D-glyceraldehyde-3-phosphate (G3P) dehydrogenase (phosphorylating) (GPDH, EC 1.2.1.12) found in liver of induced hibernating jerboa (Jaculus orientalis) was 2-3-fold lower than in the euthermic animal. However, the comparative analysis of the soluble protein fraction of these tissues by SDS-PAGE and Western blotting showed no significant changes in the intensity of the 36 kDa protein band of the GPDH subunit. After using the same purification procedure, the GPDH from liver hibernating jerboa exhibited lower values for both apparent optimal temperature and specific activity than the enzyme from the euthermic animal. Similar non-linear Arrhenius plots were obtained, but the Ea values calculated for the GPDH from hibernating tissue were higher. Although in both purified enzyme preparations four isoelectric GPDH isoforms were resolved by chromatofocusing, those of hibernating liver exhibited more acidic pI values (pI 7.3-6.1) than the hepatic isoforms of euthermic animals (pI 8.7-8.1). However, all liver GPDH isoforms exhibited similar native and subunit molecular masses and cross-reacted with an antibody raised against muscle GPDH. The comparison of the kinetic parameters of both purified preparations and the main isoforms isolated from euthermic and hibernating tissues showed the decreased catalytic efficiency of hibernating enzyme being exclusively due to a lower Vmax for both substrates G3P and NAD+. Phosphodiesterase treatment of cell-free extracts increased GPDH activity in the case of hibernating liver only. The pI of the main isoform purified from this tissue, about 6.9, changed after this treatment to an alkaline value (pI 8.44) similar to those of the euthermic GPDH isoforms. Differential ultraviolet absorption spectra of these isoforms indicated that a substance absorbing at 260 nm, that was released by the phosphodiesterase digestion, was present in the enzyme of hibernating tissue. Incubation of purified GPDH with the NO-releasing agent sodium nitroprussite produced under conditions that promote mono-ADP-ribosylation a dramatic decrease of activity (up to 60%) of both euthermic and phosphodiesterase-treated hibernating preparations but only a marginal inhibition of the hibernating enzyme. These data suggest that the liver GPDH of hibernating jerboa exhibits a posttranslational covalent modification, being probably a mono-ADP-ribosylation. The resulting inhibition of enzyme activity could contribute to the wide depression of the glycolytic metabolic flow associated with mammalian hibernation.
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PMID:Evidence for a posttranslational covalent modification of liver glyceraldehyde-3-phosphate dehydrogenase in hibernating jerboa (Jaculus orientalis). 854 42

ESTs constitute rapid and informative tools with which to study gene-expression profiles of the diverse stages of the schistosome life cycle. Following a comprehensive EST study of adult worms, analysis has now targeted the cercaria, the parasite larval form responsible for infection of the vertebrate host. Two Schistosoma mansoni cercarial cDNA libraries were examined and partial sequence obtained from 957 randomly selected clones. On the basis of database searches, 551 (57.6%) ESTs generated had no homologs in the public databases whilst 308 (32.2%) were putatively identified, totaling 859 informative ESTs. The remaining 98 (10.2%) were uninformative ESTs (ribosomal RNA and non-coding mitochondrial sequences). By clustering analysis we have identified 453 different genes. The most common sequences in both libraries represented Sm8 calcium binding protein (8% of ESTs), fructose-1,6-bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase, cytochrome oxidase subunit 1, ATP guanidine kinase and triose phosphate isomerase. One hundred and nineteen identified genes were sorted into 11 functional categories, with genes associated with energy metabolism being the most abundant (13%) and diverse. The diversity and abundance of genes associated with the transcription/translation machinery and with regulatory/signaling functions were also marked. A paramyosin transcript was identified, indicating that this gene is not exclusively expressed in adult worms and sporocysts (as had been suggested previously). The possible physiological relevance to cercariae of the presence of transcripts with homology to calcium binding proteins of the EF-hand superfamily, Gq-coupled rhodopsin photoreceptor, rod phosphodiesterase 8 subunit and peripheral-type benzodiazepine receptor is discussed.
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PMID:Analysis of the gene expression profile of Schistosoma mansoni cercariae using the expressed sequence tag approach. 1051 83

Utilisation of glucose undergoes a marked decline during erythroblastic differentiation in the chicken. Concomitantly there is a reduction in the expression of glucose transporter proteins and in the expression of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAD). GAD activity declines, after an initial rise, while the level of GAD mRNA decreases rapidly after induction of differentiation. We have employed the temperature-sensitive chicken erythroblast cell line HD3 that differentiates to the erythrocyte phenotype at 42 degrees C in the presence of inducers (hemin and butyric acid). The role of tyrosine and serine/threonine phosphorylation pathways were evaluated with the phosphatase inhibitors sodium vanadate and okadaic acid, respectively. In the presence of phosphatase inhibitors, HD3 cells underwent differentiation and increased their synthesis of hemoglobin which is a marker protein for red blood cells differentiation. The levels of both GAD mRNA and enzymatic activity were increased by phosphatase inhibitors. The role of cAMP in differentiation was also assessed. Differentiation of HD3 cells was associated with an increase in cAMP. However the phosphodiesterase inhibitor IBMX was not a good inducer of hemoglobin synthesis but did induce GAD mRNA and enzymatic activity. Together these results suggest that multiple pathways (including serine/threonine phosphorylation, tyrosine phosphorylation and elevated cAMP) are involved in the regulation of erythroblastic differentiation, hemoglobin synthesis, GAD gene expression and GAD activity in HD3 cells.
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PMID:Erythrocytic differentiation and glyceraldehyde-3-phosphate dehydrogenase expression are regulated by protein phosphorylation and cAMP in HD3 cells. 1078 56