Specificity of Cyclic GMP Activation of a Multi-substrate Cyclic Nucleotide Phosphodiesterase from Rat Liver

Abstract

Cyclic nucleotide derivatives have been used as a tool to investigate the existence of distinctive activating and hydrolytic sites 011 the phosphodiesterase from rat liver activated by cGMP (guanosine 3',5'-monophosphate). This positively cooperative enzyme was stimulated up to 30-fold by 3 pM cGMP when 3 pM CAMP (adenosine 3',5'-monophosphate) was used as substrate. All analogues were less potent activators than cGMP. Most CAMP derivatives were inactive, with two exceptions : 7-deazaadenosine 3',5'-monophosphate and 3'-amino-3'-deoxyadenosine 3',5'-monophosphate. Benzimidazole ribonucleoside 3',5'-monophosphate, where the two atoms of nitrogen of the pyrimidine ring are missing was a better stimulator than the intact purine-related cyclic derivative. When CAMP and cGMP with identical chemical ligands substituted at the same position were compared, the cGMP analogue was always the more potent activator suggesting that the activating site is sensitive to a guaninetype cyclic nucleotide structure. Degradation of the derivatives by the enzyme was measured by highperformance liquid chromatography : no relation could be established between hydrolysis and effectiveness of activation. In addition, there was no parallelism between inhibitory and activating potency for ten cyclic nucleotide derivatives. Since the chemical interactions between the analogues at the activating site on the one hand and at the catalytic sitc on the other, are different, it is proposed that the sites are distinct. Consequently, it is suggested that the enzyme operates in steps. In the first activating step, cGMP is fixed by at least two hydrogen bonds at a specific binding site of the enzyme. This is followed by a conformational change of the protein and subsequently a change of the kinetic parameters. In a rather unspecific process and in a second hydrolytic step, several purine-related cyclic nucleotides are converted to the corresponding 5' nucleotides.

Towards selective phosphodiesterase 2A (PDE2A) inhibitors: a patent review (2010 - present)

“…Although the exact mechanism of the allosteric activation by cGMP is still not completely elucidated, evidence from kinetic biochemical experiments, 15,16,17 mutagenesis studies 18 and the landmark X-ray structure by Pandit et al 19 most abundant in the brain. 22,23 This was confirmed quantitatively by comparing PDE2A mRNA expression levels between multiple human tissues.…”

A subset of GAF domains are evolutionarily conserved sodium sensors

“…At least 11 distinct families of PDE exist whose activity can be regulated by their N‐termini (Francis et al ., 2001; Houslay and Adams, 2003). Of these regulatory domains PDEs types 2, 5, 6, 10 and 11 possess GAF domains regulated by cyclic nucleotides (Erneux et al ., 1981; Mullershausen et al ., 2003; Rybalkin et al ., 2003; Zoraghi et al ., 2005; Gross‐Langenhoff et al ., 2006). The CyaB1 and CyaB2 ACs of Anabaena PCC 7120 also bind cAMP through one (CyaB1) or two (CyaB2) N‐terminal GAF domains to mediate positive feedback regulation of a carbon dioxide‐responsive AC domain (Kanacher et al ., 2002; Bruder et al ., 2005; Martinez et al ., 2005; Hammer et al ., 2006).…”

Properties and Functions of GAF Domains in Cyclic Nucleotide Phosphodiesterases and Other Proteins

“…In PDEs 2 and 5, cGMP binding to the GAF domains activates catalysis (Erneux et al, 1981;Martins et al, 1982;Okada and Asakawa, 2002;Corbin, 2003;Mullershausen et al, 2003;Rybalkin et al, 2003).…”