Phosphodiesterase activity

The cAMP formed by adenylyl cyclase (Figure 15.20) does not persist because 5 -phosphodiesterase activity prevalent in cells hydrolyzes cAMP to give 5 -AMP. Caffeine inhibits 5 -phosphodi-esterase activity. Describe the effects on glycogen phosphorylase activity that arise as a consequence of drinking lots of caffeinated coffee. 

Sircar and co-workers reported that some pyrazol-3-ones possess potent and selective inhibitory phosphodiesterase activity which is primarily responsible for their inotropic action (87JOC1724). 

Figure 18-8. Coordinated control of glycogenolysis and glycogenesis by cAMP-dependent protein kinase. The reactions that lead to glycogenolysis as a result of an increase in cAMP concentrations are shown with bold arrows, and those that are inhibited by activation of protein phosphatase-1 are shown as broken arrows. The reverse occurs when cAMP concentrations decrease as a result of phosphodiesterase activity, leading to glycogenesis.

An important observation linking Ca + to hormone action involved the definition of the intracellular targets of Ca action. The discovery of a Ca -dependent regulator of phosphodiesterase activity provided the basis for a broad understanding of how Ca and cAMP interact within cells. 

Therefore, it is currently believed that anandamide is formed from membrane phospholipids (Fig. 4) through a pathway that involves (1) a trans-acylation of the amino group of phosphatidylethanolamine with arachidonate from the sn-1 position of phosphatidylcholine and (2) a D-type phosphodiesterase activity on the resulting A-arachidonylphosphati-dylethanolamide (NAPE). Synthesis of anandamide is presumably regulated at the levels of both enzymes, the A-acyltranferase and the phospholipase D, by stimuli that raise intracellular calcium or by receptors linked with cAMP and PKA. It has been shown that anandamide is formed when neurons are depolarized and, therefore, the intracellular calcium ion levels are elevated (Cadas, 1996). 

Although caffeine is known to mobilize intracellular calcium, to inhibit phosphodiesterase activity, and to increase in vitro 5-HT and NE concentrations in the brainstem (Garrett and Griffiths 1997 Berkowitz et al. 1970 Carter et al. 1995 Solinas et al. 2002), it is now widely accepted that the mechanism of action of caffeine on wakefulness, at least at the dose range produced by voluntary caffeine intake, is via the antagonism of adenosine receptors. 

Another important group of hydrolytic enzymes are phospho- and cyclophosphodiesterases. They catalyze the hydrolysis of phospho-diester bonds and many of the most relevant biological substrates are nucleic acids. Phospholipase C and D are also important examples. Initial attempts to measure phosphodiesterase activity placed a phosphodiester between a fluorophore and a quencher and the probe was tested in vitro [146], This system was slightly modified by Caturla and used for the identification of catalysts with phosphodiesterase activity [147], More recently, Nagano and co-workers used a coumarin donor and fluorescein as a FRET... 

Fig. 6.18. Design and performance of the FRET probe CPF4 for measuring phosphodiesterase activity. Ri and R2 are phosphate and hydrogen or vice versa.

Takakusa, H., Kikuchi, K., Urano, Y., Sakamoto, S., Yamaguchi, K. and Nagano, T. (2002). Design and synthesis of an enzyme-cleavable sensor molecule for phosphodiesterase activity based on fluorescence resonance energy transfer. J. Am. Chem. Soc. 124, 1653-1657. 

There are several mechanisms involved in the vasodilator effect of flavonoids. The main mechanism seems to be related to the inhibition of protein kinase C or some of the processes activated by this protein. The inhibition of other protein kinases and cyclic nucleotide phosphodiesterase activity and blockage of calcium entry can also contribute to this effect to a greater or lesser extent (Alvarez Castro and Orallo, 2003 Herrera and others 1996). Certain flavonoids, like the flavonol myricetin, have a two-phase action on blood vessels vasoconstrictor in lowest active concentrations and vasodilator in higher concentrations (Alvarez Castro and Orallo, 2003). 

Gall 39,000 Cholera and pertussis Phosphodiesterase (activation) in rods and cones... 

Gagust 41,000 Unknown Phosphodiesterase (activation) in taste epithelium... 

M. A. Sogorb, I. Sanchez, M. Lopez-Rivadulla, V. Cespedes, E. Vilanova, EDTA-Re-sistant and Sensitive Phosphodiesterase Activities Associated with Albumin and Lipoproteins in Rabbit Serum , Drug Metab. Dispos. 1999, 27, 53-59. 

Macovschi 0, Prigent AF, Nemoz G, Pacheco FI. (1987). Effects of an extract of Ginkgo biloba on the 3, 5 -cyclic AMP phosphodiesterase activity of the brain of normal and triethyltin-intoxicated rats. J Neurochem. 49(1) 107-14. 

Mechanism of Action A positive inotropic agent that inhibits myocardial cyclic adenosine monophosphate (cAMP) phosphodiesterase activity and directly stimulates... 

Dipyridamole is a vasodilator that inhibits platelet function by inhibiting adenosine uptake and cGMP phosphodiesterase activity. Dipyridamole by itself has little or no beneficial effect. Therefore, therapeutic use of this agent is primarily in combination with aspirin to prevent cerebrovascular ischemia. It may also be used in combination with warfarin for primary prophylaxis of thromboemboli in patients with prosthetic heart valves. A combination of dipyridamole complexed with 25 mg of aspirin is now available for secondary prophylaxis of cerebrovascular disease. 

Appleman, M. M., Thompson, W. J. (1971) Multiple cyclic nucleotide phosphodiesterase activities from rat brain. Biochemistry 10(2), 311-316. 

Toeroek, K. Cowley, D.J. Brandmeier, B.D. Howell, S. Aitken, A. Tren-tham, D.R. Inhibition of calmodulin-activated smooth-muscle myosin light-chain kinase by calmodulin-binding peptides and fluorescent (phosphodiesterase-activating) calmodulin derivatives. Biochemistry, 37, 6188-6198 (1998)... 

The conclusion of Bernardi and Griffe (3) that the phosphodiesterase activity of acid DNase is an intrinsic property of the enzyme molecule has been recently challenged by Slor (46, 58), Swenson and Hodes (54), and Slor and Hodes (55), who claimed to have obtained a separation of the two activities. In fact, none of the reported results proves an actual separation of the two activities and constitutes an acceptable evidence against the two activities being carried by the same protein molecule. Some data suggest, however, that the phosphodiesterase activity may be inactivated preferentially by some treatments. In connection with the phosphodiesterase activity of acid DNase, see also Tables I and II in reference (56) and the related discussion (56a). 

Scheme 12.3 Adenylate cyclase/phosphodiesterase activity by cyclodextrin complexes.2...

The lipophilic zinc(II) complex 11 (0.25-1.0 mM) with 10 mM Triton X-100 also showed phosphodiesterase activity in BNP (5-10 mM) hydrolysis at 35°C with similar kinetic behavior to that exhibited in the NA and TNP hydrolysis. The rate-pH profile curve disclosed an inflection point at pH 8.3 with 1.0 mM 11, indicating that the kinetically active species was again the zinc(II)-bound OH- complex lib (Scheme 9). The second-order rate constant BNP (see Eq. [9]) for the hydrolysis of BNP promoted by lib was 4.3 X 10-4 M-1 sec-1 at 35°C and pH 10.2 (20 mM CAPS buffer) with I = 0.10 (NaN03). The fcBNP value for lib with 10 mM Triton X-100 is only approximately 20 times larger than the reported kBNP value for 2b (2.1 X 10-5 M-1 sec-1) [9] under the same conditions. The rate enhancement (i.e., 20 times) by the metallomicelles is not as dramatic as that for the more lipophilic TNP (290 times) and NA (50 times), because the less lipophilic nature of BNP relative to that of TNP and NA would result in smaller partition into the metallomicelles. 

Effects of Dopaminergic Antagonists on Calmodulin-stimulated Phosphodiesterase Activity... 

Both calmodulin and a calcium- and calmodulin-dependent phosphodiesterase activity occur in dispersed bovine parathyroid cells (20.). Some of the phenothiazines blocking the dopamine receptor in this tissue inhibit the interaction between calmodulin and the calmodulin-dependent phosphodiesterase. A series of experiments have tested the possibility that calmodulin might play a role in the effects of these agents on the parathyroid gland. The dopamine antagonists examined were weak inhibitors of calmodulin-stimulated phosphodiesterase activity (Table II). 

The stereospecific dopaminergic antagonists (-) and (+) butaclamol and cis- and trans-flupenthixol were of equal potency in their effects on calmodulin-dependent phosphodiesterase activity. This contrasts with the marked difference in the potency of each pair of stereoisomers as dopamine antagonists. It appears unlikely that the calmodulin-dependent phosphodiesterase activity (and by inference calmodulin) plays any role in the actions of dopaminergic antagonists upon the bovine parathyroid gland. 

The role of cyclic AMP as modulator of prolactin secretion was first suggested by the finding of a stimulatory effect of cyclic AMP derivatives (17-22) and inhibitors of cyclic nucleotide phosphodiesterase activity such as theophylline and IBMX (22-26) on the secretion of this hormone. More convincing evidence supporting a role of cyclic AMP in the action of dopamine on prolactin secretion had to be obtained, however, by measurement of adenohypophysial adenylate cyclase activity or cyclic AMP accumulation under the influence of the catecholamine. As illustrated in Fig. 1, addition of 100 nM dopamine to male rat hemipituitaries led to a rapid inhibition of cyclic AMP accumulation, a maximal effect (30% inhibition) being already obtained 5 min after addition of the catecholamine. Thus, while dopamine is well known to stimulate adenylate cyclase activity in the striatum (27, 28), its effect at the adenohypophysial level in intact cells is inhibitory. Dopamine has also been found to exert parallel inhibitory effects on cyclic AMP levels and prolactin release in ovine adenohypophysial cells in culture (29) and purified rat mammotrophs (30). Using paired hemipituitaries obtained from female rats, Ray and Wallis (22) have found a rapid inhibitory effect of dopamine on cyclic AMP accumulation to approximately 75% of control. 

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