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 new assay for cyclic nucleotide phosphodiesterase activity by high-performance liquid chromatography with on-line radiochemical detection has been developed. The method is based on the measurement of 3H-labeled nucleoside monophosphates formed from cyclic nucleotides by the action of 3',5'-cyclic-nucleotide phosphodiesterase (PDE). The reaction products are determined from the incubation mixture after removal of the protein by injection of an aliquot into the liquid chromatograph. The detection limit with counting efficiency of 30% is 20 fmol of 3H-labeled product, which makes the method suitable for detection of low PDE activities.
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PMID:Determination of cyclic nucleotide phosphodiesterase activity by high-performance liquid chromatography. 244 23

Adenosine 3',5'-cyclic monophosphate (cAMP) concentration and 3',5'-cyclic-nucleotide phosphodiesterase (PDE) activity were measured in skeletal muscle, heart, and liver of rats exposed to 1, 3, 5, and 7 days of cold. Cyclic nucleotide concentration increased in fast-twitch red muscle at the same time that PDE activity was decreasing. Nucleotide concentration and enzyme activity of slow-twitch red muscle were not altered by the cold exposure. The PDE activity of fast-twitch white muscle was elevated approximately 50% above control after 1 and 3 days of cold exposure. By the 5th day in the cold, white muscle PDE activity had returned to control levels and remained there through the 7th day of experimentation. cAMP concentration in hearts of cold-exposed rats was significantly (P less than 0.01) elevated above control at all time points measured. Myocardial PDE activity was elevated above control (P less than 0.05) at 1 and 3 days of cold exposure but returned to control levels by the 5th day in the cold. Hepatic cAMP and PDE activity were elevated above control at all time points analyzed. These data suggest that changes in cyclic nucleotide metabolism play a role in attaining homeostasis during acute cold exposure.
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PMID:Effect of cold exposure on liver and muscle cAMP content and cAMP phosphodiesterase activity. 298 94

Several steps of purification (octyl-Sepharose chromatography, Blue Sepharose 6B chromatography and sucrose density gradient centrifugation) led to a highly purified aggregate of the enzymes, 3',5'-cyclic-nucleotide phosphodiesterase (PDE) and nucleotidase. The purified enzyme aggregate showed an S value of 7.3 (SE +/- 0.3, n = 10). Further analysis by SDS-polyacrylamide gel electrophoresis (PAGE) revealed two proteins near 67 and 60 kDa. Dissociation of the 7.3 S enzyme aggregate showed a 3.6 S PDE form and a nucleotidase form at 4.2 S. Additionally, higher S value forms of the nucleotidase up to 17 S have been observed. Apparently, they had formed by self-association. SDS-PAGE of the 17 S nucleotidase form showed only one band at 67 kDa. This was taken as evidence for the homogeneity of the 17 S nucleotidase form and the self-association of the nucleotidase after dissociation from the 7.3 S enzyme aggregate. Furthermore, from this it could be concluded that the 67 kDa protein of the 7.3 S enzyme aggregate should be identified with the nucleotidase, and thus the 60 kDa band represents the PDE.
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PMID:Purification of an enzyme aggregate containing 3',5'-cyclic-nucleotide phosphodiesterase and nucleotidase. 301 49

A procedure for nondenaturing immunopurification of bovine calmodulin-dependent 3',5'-cyclic-nucleotide phosphodiesterase (3',5'-cyclic-nucleotide 5'-nucleotidohydrolase, EC 3.1.4.17) is described that utilizes chromatography on a conformation-specific monoclonal antibody column. Hybridomas derived from spleen cells of mice immunized with Ca(2+)/calmodulin/phosphodiesterase were screened for antiphosphodiesterase antibody production. A stable cell line was established that secretes a monoclonal antibody that binds to the Ca(2+)/calmodulin/enzyme complex with an approximate K(d) of 10(-9) M. The dissociation constant was increased by two orders of magnitude when calmodulin interaction with the enzyme was inhibited by Ca(2+) chelation. This differential reactivity was utilized for affinity chromatography of heart and brain phosphodiesterases on monoclonal antibody columns. Highly purified phosphodiesterases were eluted in good yield with buffer containing EGTA. The immunopurified enzymes from heart and brain exhibited specific activities of approximately 300 units/mg when assayed at millimolar concentrations of cGMP or cAMP. Calmodulin stimulated both enzymes 10- to 15-fold over basal activity under these conditions. However, analysis of the two preparations by NaDodSO(4)/polyacrylamide gel electrophoresis revealed an apparent subunit of M(r) 61,000 for the brain enzyme, in contrast to the M(r) 59,000 cardiac subunit. The observed difference was not an artifact of tissue homogenization because both forms were detected after purification from mixed-tissue homogenates. These results suggest that mild, biospecific elution from a conformation-specific monoclonal antibody column may be a general technique applicable to the rapid isolation of proteins whose antigenic determinants can be altered with specific ligands.
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PMID:Purification of two calcium/calmodulin-dependent forms of cyclic nucleotide phosphodiesterase by using conformation-specific monoclonal antibody chromatography. 628 44

The interaction between selective inhibitors of 3',5'-cyclic-nucleotide phosphodiesterase (PDE) III (cyclic GMP inhibited phosphodiesterase) and selective inhibitors of PDE IV (Ro 20-1724 inhibited phosphodiesterase) to attenuate fetal bovine serum-stimulated incorporation of [3H]thymidine into DNA and cell proliferation was studied in a line (A10) of vascular smooth muscle cells (VSMC). The nonselective PDE inhibitors 3-isobutyl-1-methylxanthine (IBMX) and papaverine attenuated DNA synthesis with EC50 values (16 and 18 microM, respectively) in the same range as their published IC50 values (2-50 and 2-25 microM, respectively) as PDE inhibitors. The selective PDE III inhibitors CI-930 and cilostamide used alone attenuated DNA synthesis with EC50 values (> 300 and 5.3 microM, respectively) that were much higher than published IC50 values (0.15-0.46 and 0.005-0.064 microM, respectively) for inhibition of PDE III. In the presence of the PDE IV inhibitor rolipram (10 microM), their EC50 values were shifted (0.66 and 0.16 microM, respectively) much closer to their respective IC50 values. When the selective PDE IV inhibitors rolipram and Ro 20-1724 were used alone, they attenuated DNA synthesis with EC50 values (111 and > 100 microM, respectively) much higher than their IC50 values (0.6-2.6 and 2-13 microM, respectively) as inhibitors of PDE IV, but 10 microM CI-930 (PDE III inhibitor) shifted their EC50 values (0.56 and 1.5 microM, respectively) much closer to their IC50 values. In experiments that assessed VSMC proliferation using the MTT [3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] method, IBMX and papaverine attenuated proliferation with EC50 values (27 and 58 microM, respectively) close to their IC50 values. CI-930 and cilostamide used alone did not cause 50% attenuation of proliferation at the highest concentrations tested (100 and 10 microM, respectively). In the presence of 5 microM rolipram, however, their effects were enhanced greatly with EC50 values (0.86 and 0.23 microM, respectively) that were close to their IC50 values as PDE III inhibitors. Similarly, rolipram and Ro 20-1724 attenuated VSMC proliferation with EC50 values close to their IC50 values in the presence (2.1 and 4.6 microM, respectively) but not in the absence (> 100 and > 10 microM, respectively) of 2 microM CI-930. The interactions between PDE III inhibitors and PDE IV inhibitors to attenuate DNA synthesis and VSMC proliferation were synergistic as determined by the combination index. The data demonstrate that the synergistic interactions that attenuate incorporation of [3H]thymidine into DNA are accompanied by synergistic attenuations of VSMC division.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Synergistic interactions between selective pharmacological inhibitors of phosphodiesterase isozyme families PDE III and PDE IV to attenuate proliferation of rat vascular smooth muscle cells. 752 42