scholarly journals The metabolism of cyclic nucleotides in the guinea-pig pancreas. Cyclic AMP phosphodiesterase and cyclic GMP phosphodiesterase

1980 ◽  
Vol 186 (2) ◽  
pp. 491-498 ◽  
Author(s):  
Patricia Methven ◽  
Marius Lemon ◽  
Kanti Bhoola
Keyword(s):  
Cyclic Amp ◽  
Guinea Pig ◽  
Cyclic Gmp ◽  
Cyclic Nucleotide ◽  
Metal Ion ◽  
Gel Filtration ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Physiological Control ◽  
Cyclic Amp Phosphodiesterase ◽  
Stimulus Secretion Coupling

Both cyclic AMP phosphodiesterase and cyclic GMP phosphodiesterase were recovered mainly from the supernatant fractions of guinea-pig pancreas, but a higher proportion of the activity of the former was associated with the pellet fractions. The activities in the supernatant were not separated by gel filtration, but were clearly separated by subsequent chromatography on an anion-exchange resin. The activities of cyclic AMP phosphodiesterase and cyclic GMP phosphodiesterase had high-affinity (Km 6.5±1.1μm and 31.9±3.9μm respectively) and low-affinity (Km 0.56±0.05mm and 0.32±0.03mm respectively) components. The activity of neither enzyme was affected by the pancreatic secretogens, cholecystokinin-pancreozymin, secretin and carbachol. Removal of ions by gel filtration resulted in a marked reduction in cyclic nucleotide phosphodiesterase activity, which could be restored by addition of Mg2+. Mn2+ (3mm) was as effective as Mg2+ (3mm) in the case of cyclic AMP phosphodiesterase, but was less than half as effective in the case of cyclic GMP phosphodiesterase. The metal-ion chelators, EDTA and EGTA, also decreased activity. Ca2+ (1mm) did not affect the activity of cyclic nucleotide phosphodiesterase when the concentration of Mg2+ was 3mm. At concentrations of Mg2+ between 0.1 and 1mm, 1mm-Ca2+ was activatory, and at concentrations of Mg2+ below 0.1mm, 1mm-Ca2+ was inhibitory. These results are discussed in terms of the possible significance of cyclic nucleotide phosphodiesterase in the physiological control of cyclic nucleotide concentrations during stimulus–secretion coupling.

scholarly journals The identification of a new cyclic nucleotide phosphodiesterase activity in human and guinea-pig cardiac ventricle. Implications for the mechanism of action of selective phosphodiesterase inhibitors

1987 ◽  
Vol 241 (2) ◽  
pp. 535-541 ◽  
Author(s):  
M L Reeves ◽  
B K Leigh ◽  
P J England
Keyword(s):  
Cyclic Amp ◽  
Guinea Pig ◽  
Cardiac Muscle ◽  
Cyclic Gmp ◽  
Cyclic Nucleotide ◽  
Gel Filtration ◽  
Phosphodiesterase Inhibitors ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Kinetic Properties ◽  
Cardiac Ventricle

Four cyclic nucleotide phosphodiesterase (PDE) activities were separated from low-speed supernatants of homogenates of human cardiac ventricle by DEAE-Sepharose chromatography, and designated PDE I-PDE IV in order of elution with an increasing salt gradient. PDE I was a Ca2+/calmodulin-stimulated activity, and PDE II was an activity with a high Km for cyclic AMP which was stimulated by low concentrations of cyclic GMP. Human ventricle PDE III had Km values of 0.14 microM (cyclic AMP) and 4 microM (cyclic GMP), and showed simple Michaelis-Menten kinetics with both substrates. PDE IV is a previously unrecognized activity in cardiac muscle, the human enzyme having Km values of 2 microM (cyclic AMP) and 50 microM (cyclic GMP). PDE III and PDE IV were not activated by cyclic nucleotides or calmodulin. Four PDE activities were also isolated from guinea-pig ventricle, and had very similar kinetic properties. By gel filtration, the Mr of PDE III was 60,000, and that of PDE IV 45,000. The drug SK&F 94120 selectively and competitively inhibited PDE III with a Ki value of 0.8 microM (human), showing simple hyperbolic inhibition kinetics. Rolipram (Schering ZK 62711) and Ro 20-1724 (Roche), which have previously been reported to inhibit PDE III-like activities strongly, were shown to be weak inhibitors of human and guinea-pig PDE III enzymes (Ki values greater than 25 microM), but potent inhibitors of PDE IV [Ki values 2.4 microM (Rolipram) and 3.1 microM (Ro 20-1724) with human PDE IV]. The inhibition in all cases demonstrated simple hyperbolic competition. These observations suggest that the previously reported complex inhibition of PDE III-type activities from cardiac muscle was caused by incomplete separation of the PDE III from other enzymes, particularly PDE IV.

scholarly journals Cyclic nucleotide phosphodiesterase activities in Neurospora crassa

1982 ◽  
Vol 203 (3) ◽  
pp. 611-616 ◽  
Author(s):  
M T Téllez-I ñón ◽  
G C Glikin ◽  
H N Torres
Keyword(s):  
Cyclic Amp ◽  
Neurospora Crassa ◽  
Cyclic Gmp ◽  
Cyclic Nucleotide ◽  
Gradient Centrifugation ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Sucrose Density Gradient Centrifugation ◽  
Molecular Weights

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.

scholarly journals Cyclic AMP phosphodiesterase and cyclic GMP phosphodiesterase activities of rat mammary tissue

1984 ◽  
Vol 219 (3) ◽  
pp. 801-809 ◽  
Author(s):  
I Mullaney ◽  
R A Clegg
Keyword(s):  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Cyclic Nucleotide ◽  
Deae Cellulose ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Mammary Tissue ◽  
Phosphodiesterase Activity ◽  
High Affinity ◽  
Low Concentrations ◽  
Maximal Activation

Cyclic nucleotide phosphodiesterase activity in mammary tissue from rats in midlactation was resolved by DEAE-cellulose chromatography into three functionally distinct fractions: a Ca2+/calmodulin-stimulated cyclic GMP phosphodiesterase, a cyclic GMP-stimulated low-affinity cyclic nucleotide phosphodiesterase, and a high-affinity cyclic AMP-specific phosphodiesterase. The absolute activities and relative proportions of high- and low-affinity enzymes resemble those found, for example, in liver, as distinct from those in excitable tissues. Three functional characteristics are described which are peculiar to mammary-tissue phosphodiesterases. Firstly, the concentration of free Ca2+ required to achieve half-maximal activation of the Ca2+/calmodulin-stimulated phosphodiesterase is somewhat higher than for the analogous enzyme in other tissues; secondly, the activity of this enzyme towards cyclic AMP relative to that towards cyclic GMP is unusually low, and thirdly, the low-affinity cyclic nucleotide phosphodiesterase is inhibited by low concentrations of free Ca2+.

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