Binding of cyclic nucleotides to phosphodiesterase 10A and 11A GAF domains does not stimulate catalytic activity.

To date eleven human PDE (3',5'-cyclic nucleotide phosphodiesterase) families have been identified. Of these, five families contain non-catalytic tandem GAF (cGMP-specific and -stimulated phosphodiesterases, Anabaena adenylate cyclases and Escherichia coli FhlA) domains, GAFa and GAFb, in the N-terminal part of the enzyme. For PDE2A, PDE5A and PDE6 the GAF domains have been shown to bind cGMP with high affinity. For PDE2A and PDE5A this ligand binding has been shown to stimulate the catalytic activity of the enzyme. PDE10A and PDE11A are the two most recently described PDEs and it has been suggested that their GAF domains bind to cAMP and cGMP respectively. We have developed a scintillation proximity-based assay to directly measure cyclic nucleotide binding to the PDE2A, PDE10A and PDE11A GAF domains, and in the present study we demonstrate binding of cyclic nucleotides to the PDE10A and PDE11A GAF domains. We show that these non-catalytic sites bind cAMP and cGMP respectively with much higher affinity than has previously been suggested using indirect assessment of the interaction. The GAFb domain of PDE10A binds cAMP with a Kd of 48 nM and the GAFa domain of PDE11A binds cGMP with a Kd of 110 nM. The effect of cyclic nucleotides binding to the GAF domains on the enzyme activity was investigated through the use of modified cyclic nucleotides. In contrast with other GAF domain-containing PDEs, and with what has previously been predicted, ligand binding to the GAF domains of PDE10A and PDE11A does not stimulate catalytic activity.