Characterization of Cyclic Nucleotide Phosphodiesterases of Rat Tissues

Thompson, W. J.; Appleman, M. Michael

doi:10.1016/s0021-9258(18)62207-0

Cited by 293 publications

(28 citation statements)

References 16 publications

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“…The specific activity was some five to sixfold less than in the parent synaptosome fraction. However, there are some reports in the literature (3,8,13,15,38,44,46) describing the existence of multiple forms of the cyclic nucleotide-phosphodiesterase in brain depending on specific cation activation and on substrate specificity, so one cannot make a valid comparison between the specific activities in the synaptosome and PSD until the exact nature of the brain phosphodiesterase activities is known. Because the PSD preparation contains both a cyclic nucleotide phosphodiesterase activity and calmodulin, it was assumed that this preparation contained a calmodulin-activatable cyclic nucleotide phosphodiesterase activity.…”

Section: Resultsmentioning

confidence: 99%

“…Because several multiple forms of the phosphodiesterase have been described in brain (3,8,15,38,44,46) it was of interest to initially characterize by gel filtration the enzymatic activity in the sonicated supernatant fraction from the PSD . To obtain enough activity for gel filtration it was necessary to use a higher concentration (13 mg PSD protein/5 ml) for the sonication; under these conditions only 20-46% of the initial activity was solubilized .…”

Section: Resultsmentioning

confidence: 99%

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Function of calmodulin in postsynaptic densities. I. Presence of a calmodulin-activatable cyclic nucleotide phosphodiesterase activity.

Grab

Carlin

Siekevitz

1981

The Journal of Cell Biology

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The postsynaptic density (PSD) fraction from canine cerebral cortex was found to contain an endogenous cyclic nucleotide-phosphodiesterase activity that was dependent on Mn 2+ and/or Mg" but not on Ca t + . Maximal activity was obtained at 1 gm Mn" . This cyclic nucleotide phosphodiesterase activity was not decreased upon removal of the calmodulin from the PSD fraction, nor was it increased by the addition of calmodulin to a postsynaptic density fraction deficient in calmodulin . The enzymatic activity could be extracted by sonication, with the soluble enzyme having properties similar to those found in the native structure. Two peaks of cyclic nucleotide phosphodiesterase activities could be obtained after S-300 Sephacryl column chromatography of this soluble fraction : fraction I (excluded peak) and fraction II (215,000 mol wt) . The fraction I activity preferred cyclic AMP over cyclic GMP and was not activated by calmodulin . The fraction II activity had an approximately fourfold lower K,n for cyclic GMP over cyclic AMP. This fraction II activity was activatable by calmodulin, which increased the Vmsx and decreased the Km in the case of both cyclic nucleotides. We conclude that two activities are present in the PSD, one activatable, and one not activatable, by Calmodulin .The postsynaptic density (PSD) is a prominent structure, of unknown function, in central nervous system synapses (1). As observed by electron microscopy, the PSD appears as a densely staining thickening which lies along the cytoplasmic side of the postsynaptic membrane (1) . Recently, we have isolated from canine cerebral cortex a PSD fraction that has been found to contain some 10 major and at least 20 minor proteins (10). Of the enzymatic activities tested, there are no Mg" or Cat +-ATPase (10) nor adenylate cyclase (18) activities, and very small amounts of cytochrome c oxidase (10) or 5'-nucleotidase (10) activities. However, two protein kinase activities are present, one activatable by cyclic AMP (4, 45) and one activatable by Ca" and calmodulin (7,18,20,21) ; the protein substrates, proteins Ia and Ib, which are phosphorylated by the former kinase (4, 45) and the protein substrates, the major 51,000 Mr protein and a 62,000 M, protein, which are phosphorylated by the latter kinase (7,18,20,21) are also found in the PSD preparation .Immunochemistry has been used to confirm the presence of proteins Ia and Ib in the PSD (6) . Other proteins identified to be intrinsic constituents are actin (5, 27), the alpha and beta subunits of tubulin (5,27,35,36,48) and a major M, = 51,000

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Function of calmodulin in postsynaptic densities. I. Presence of a calmodulin-activatable cyclic nucleotide phosphodiesterase activity.

Grab

Carlin

Siekevitz

1981

The Journal of Cell Biology

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“…Phosphodiesterase activity was determined according to the method of Thompson and Appleman (1971). Aliquots of particulate or soluble fraction (0.1-0.3 mg/ ml) were incubated in a 200-ml final volume, for 5 min at 20 C, with PDE buffer: 50 mM Tris-HCl, pH 8.0, 5 mM MgCl 2 , 3 mM b-mercaptoethanol, 0.2 mM cAMP, 0.05 mCi (1.85 kBq) of [ 3 H]cAMP.…”

Section: Measurements Of Phosphodiesterase Activitymentioning

confidence: 99%

Identification and partial characterization of cAMP-phosphodiesterases in the ciliate Euplotes raikovi

Apone

Pretoro

Vallesi

2004

European Journal of Protistology

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“…Phosphodiesterase activity was determined according to the method of Thompson and Appleman [19] in total cell lysates prepared as previously described for measurements of the adenylate cyclase activity. These lysates were incubated in a 200-l final volume, for 5 min at 20°C, with buffer (50 mM Tris-HCl, pH 8.0, 5 mM MgCl 2 ), 3 mM ␤-mercaptoethanol, and 0.2 mM cAMP containing 0.1 Ci [ 3 H]cAMP.…”

Section: Measurements Of Phosphodiesterase Activitymentioning

confidence: 99%

Cross-talk between the autocrine (mitogenic) pheromone loop of the ciliate euplotes raikovi and the intracellular cyclic AMP concentration

Apone

Vallesi

Pretoro

et al. 2003

Experimental Cell Research

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Characterization of Cyclic Nucleotide Phosphodiesterases of Rat Tissues

Cited by 293 publications

References 16 publications

Function of calmodulin in postsynaptic densities. I. Presence of a calmodulin-activatable cyclic nucleotide phosphodiesterase activity.

Function of calmodulin in postsynaptic densities. I. Presence of a calmodulin-activatable cyclic nucleotide phosphodiesterase activity.

Identification and partial characterization of cAMP-phosphodiesterases in the ciliate Euplotes raikovi

Cross-talk between the autocrine (mitogenic) pheromone loop of the ciliate euplotes raikovi and the intracellular cyclic AMP concentration

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