scholarly journals Characterization of a calmodulin-dependent high-affinity cyclic AMP and cyclic GMP phosphodiesterase from male mouse germ cells

1984 ◽  
Vol 217 (3) ◽  
pp. 693-700 ◽  
Author(s):  
R Geremia ◽  
P Rossi ◽  
D Mocini ◽  
R Pezzotti ◽  
M Conti
Keyword(s):  
Cyclic Amp ◽  
Germ Cells ◽  
Cyclic Gmp ◽  
Salt Concentration ◽  
Male Mouse ◽  
Competitive Inhibition ◽  
Sedimentation Coefficient ◽  
Phosphodiesterase Activity ◽  
Thermal Stabilities ◽  
High Affinity

Two cyclic nucleotide phosphodiesterase activities were separated by ion-exchange chromatography of cytosol from male mouse germ cells. A form eluted at low salt concentration showed high affinity (Km congruent to 2 microM) and low affinity (Km congruent to 20 microM) for cyclic AMP, and high affinity (Km congruent to 3.5 microM) for cyclic GMP. A second form, eluted at high salt concentration, showed high affinity (Km congruent to 5 microM) for cyclic AMP and was similar to a phosphodiesterase activity described in rat germ cells. The present study was performed to characterize the first form, which represents most of the phosphodiesterase activity in mouse germ cells. The enzyme was sensitive to Ca2+ and calmodulin stimulation, which increased its activity 3-4-fold. Calmodulin stimulation depended on direct interaction of the activator with the enzyme, as indicated by the reversible changes in the chromatographic elution pattern in the presence of Ca2+, as well as by the increase in the sedimentation coefficient in the presence of calmodulin. Reciprocal inhibition kinetics between cyclic AMP and cyclic GMP for the calmodulin-dependent form demonstrated a non-competitive inhibition between the two substrates, suggesting the presence of separate catalytic sites. This is in agreement with kinetic parameters and different thermal stabilities of cyclic AMP- and cyclic GMP-hydrolysing activities. Furthermore, the relevant change in s value, depending on the absence or presence of Ca2+ and calmodulin, suggested that the enzyme is composed of subunits, which aggregate in the presence of the activator. A model for catalytic site composition and reciprocal interaction is also proposed.

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+.

Cloning and characterization of the low-affinity cyclic AMP phosphodiesterase gene of Saccharomyces cerevisiae

1987 ◽  
Vol 7 (10) ◽  
pp. 3629-3636
Author(s):  
J Nikawa ◽  
P Sass ◽  
M Wigler
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cyclic Amp ◽  
Copy Number ◽  
Growth Arrest ◽  
Yeast Cells ◽  
Phosphodiesterase Activity ◽  
Camp Phosphodiesterase ◽  
High Affinity ◽  
Cyclic Amp Phosphodiesterase

Saccharomyces cerevisiae contains two genes which encode cyclic AMP (cAMP) phosphodiesterase. We previously isolated and characterized PDE2, which encodes a high-affinity cAMP phosphodiesterase. We have now isolated the PDE1 gene of S. cerevisiae, which encodes a low-affinity cAMP phosphodiesterase. These two genes represent highly divergent branches in the evolution of phosphodiesterases. High-copy-number plasmids containing either PDE1 or PDE2 can reverse the growth arrest defects of yeast cells carrying the RAS2(Val-19) mutation. PDE1 and PDE2 appear to account for the aggregate cAMP phosphodiesterase activity of S. cerevisiae. Disruption of both PDE genes results in a phenotype which resembles that induced by the RAS2(Val-19) mutation. pde1- pde2- ras1- ras2- cells are viable.

scholarly journals Evidence for a messenger function of cyclic GMP during phosphodiesterase induction in Dictyostelium discoideum

1982 ◽  
Vol 152 (1) ◽  
pp. 232-238
Author(s):  
P J van Haastert ◽  
F J Pasveer ◽  
R C van der Meer ◽  
P R van der Heijden ◽  
H van Walsum ◽  
...  
Keyword(s):  
Computer Simulation ◽  
Folic Acid ◽  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Cyclic Gmp ◽  
Close Correlation ◽  
Phosphodiesterase Activity ◽  
Transient Elevation ◽  
Lines Of Evidence ◽  
Stimulation Of

Chemotactic stimulation of vegetative or aggregative Dictyostelium discoideum cells induced a transient elevation of cyclic GMP levels. The addition of chemoattractants to postvegetative cells by pulsing induced phosphodiesterase activity. The following lines of evidence suggest a messenger function for cyclic GMP in the induction of phosphodiesterase: (i) Folic acid and cyclic AMP increased cyclic GMP levels and induced phosphodiesterase activity. (ii) Cyclic AMP induced both cyclic GMP accumulation and phosphodiesterase activity by binding to a rate receptor. (iii) The effects of chemical modification of cyclic AMP or folic acid on cyclic GMP accumulation and phosphodiesterase induction were closely correlated. (iv) A close correlation existed between the increase of cyclic GMP levels and the amount of phosphodiesterase induced, independent of the type of chemoattractant by which this cyclic GMP accumulation was produced. (v) Computer simulation of cyclic GMP binding to intracellular cyclic GMP-binding proteins indicates that half-maximal occupation by cyclic GMP required the same chemoattractant concentration as did half-maximal phosphodiesterase induction.

scholarly journals Analysis of a Dictyostelium chemotaxis mutant with altered chemoattractant binding

1988 ◽  
Vol 91 (4) ◽  
pp. 479-489 ◽  
Author(s):  
J.E. Segall ◽  
A.A. Bominaar ◽  
E. Wallraff ◽  
R.J. De Wit
Keyword(s):  
Linkage Group ◽  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Binding Sites ◽  
Group Iv ◽  
Cell Area ◽  
Wild Type ◽  
High Affinity ◽  
In The Wild ◽  
Gmp Production

A Dictyostelium discoideum mutant defective in folate chemotaxis has been analysed using biochemical, behavioural, and genetic methods. A subset of the cell-surface folate binding sites appeared to be locked in a high-affinity state from which folate dissociated extremely slowly. Changes in cell area and motility induced by step increases in folate required 10- to 100-fold higher concentrations than in the wild type. Folate-stimulated cyclic GMP production was also altered. Chemotactic responses to cyclic AMP as well as cyclic AMP-stimulated cyclic GMP production were normal. The mutation responsible for the chemotaxis defect, termed folA1000, was localized to linkage group IV. The alterations in folate binding and sensitivity to folate co-localized with the folA1000 mutation. We conclude that the folA1000 mutation arrests the folate chemotaxis receptor in a high affinity state that can only poorly transduce folate binding into chemotactic responses.

STUDIES ON THE MECHANISMS OF SOMATOSTATIN ACTION ON INSULIN RELEASE

1977 ◽  
Vol 85 (2) ◽  
pp. 379-388 ◽  
Author(s):  
A Claro ◽  
V. Grill ◽  
S. Efendić ◽  
R. Luft
Keyword(s):  
Cyclic Amp ◽  
Insulin Release ◽  
Competitive Inhibition ◽  
Phosphodiesterase Inhibitor ◽  
Adenyl Cyclase ◽  
Phosphodiesterase Activity ◽  
High Concentration ◽  
Isolated Islets Of Langerhans ◽  
Cyclic Amp Accumulation ◽  
Effect Of Glucose

ABSTRACT The effects of somatostatin on insulin release and cyclic AMP metabolism were studied in collagenase-isolated islets of Langerhans from the rat. Concentrations from 500 to 2000 ng/ml significantly inhibited glucose stimulated insulin release, while 100 and 200 ng/ml were ineffective. Somatostatin (2000 ng/ml) inhibited insulin release and [3H]-cyclic AMP accumulation induced by 16.7 mm glucose after 10 and 30 min of incubation. In dose-response studies, the inhibition by somatostatin of the effect of glucose on [3H] cyclic AMP and insulin release could be overcome by a high concentration of the hexose (44.9 mM), suggesting competitive inhibition. In the absence of glucose, somatostatin inhibited [3H] cyclic AMP accumulation induced by the phosphodiesterase inhibitor, IBMX, while no inhibition was seen, again in the absence of hexose, when the [3H] cyclic AMP levels had been raised by the adenyl cyclase stimulator, cholera toxin. Somatostatin did not affect phosphodiesterase activity when added to islet homogenates, but preincubation of the islets with the peptide before homogenization decreased the activity by about 30 %. It is suggested that somatostatin-induced inhibition of insulin release is, at least partially, mediated by cyclic AMP, probably through an action on islet adenyl cyclase.

scholarly journals Measurements of cyclic AMP and cyclic GMP phosphodiesterase activity in isolated tubular segments

1980 ◽  
Vol 18 (4) ◽  
pp. 512-518 ◽  
Author(s):  
Brian A. Jackson ◽  
Richard M. Edwards ◽  
Thomas P. Dousa ◽  
Denise M. Heublein ◽  
Julie L. Braun
Keyword(s):  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Phosphodiesterase Activity

scholarly journals Cyclic nucleotide phosphodiesterase of rat pancreatic islets. Effects of Ca2+, calmodulin and trifluoperazine

1981 ◽  
Vol 197 (2) ◽  
pp. 459-464 ◽  
Author(s):  
M C Sugden ◽  
S J Ashcroft
Keyword(s):  
Cyclic Amp ◽  
Pancreatic Islets ◽  
Islets Of Langerhans ◽  
Cyclic Gmp ◽  
Cyclic Nucleotide ◽  
Specific Inhibitor ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Phosphodiesterase Activity ◽  
Rat Pancreatic Islets ◽  
Rat Islets Of Langerhans

Cyclic nucleotide phosphodiesterase activity towards cyclic AMP and cyclic GMP was studied in extracts of rat islets of Langerhans. Biphasic Eadie plots [Eadie (1942) J. Biol. Chem. 146, 85-93] were obtained with either substrate suggesting the presence of both ‘high’- and ‘low’-Km components. The apparent Km values were 6.2 +/- 0.5 (n = 8) microM and 103.4 +/- 13.5 (6) microM for cyclic AMP and 3.6 +/- 0.3 (12) microM and 61.4 +/- 7.5 (13) microM for cyclic GMP. With cyclic AMP as substrate, phosphodeisterase activity was increased by calmodulin and Ca2+ and decreased by trifluoperazine, a specific inhibitor of calmodulin. With cyclic GMP as substrate, phosphodiesterase activity was decreased by omission of Ca2+ or addition of trifluoperazine. Addition of exogenous calmodulin had no effect on activity. The data suggest that Ca2+ may influence the islet content of cyclic AMP and cyclic GMP via effects on calmodulin-dependent cyclic nucleotide phosphodiesterase(s).

Sign in / Sign up

Export Citation Format

Share Document