Characterization of cyclic nucleotide phosphodiesterases with cyclic AMP analogs: topology of the catalytic sites and comparison with other cyclic AMP-binding proteins.
To define essential interactions of cAMP with the catalytic sites of cyclic nucleotide phosphodiesterases (PDEs) and to begin to map the topology of the sites, we have tested a series of cAMP analogs as competitive inhibitors of the PDEs that hydrolyze cAMP with high efficiency (PDE1, PDE2, PDE3, and PDE4). Comparisons of IC50 values, relative to cAMP, were used to predict which functional groups on cAMP interact with each isozyme. Common to all PDEs tested, except for the calcium/calmodulin-dependent PDE (CaM-PDE, PDE1), is an interaction at the N1-position of cAMP and a distinct lack of binding to the 2'-hydroxyl group of the ribose moiety. Only the cGMP-stimulated (PDE2) and cAMP-specific (PDE4) PDEs appear to interact strongly at the N7-position. The cGMP-inhibited PDE (cGI-PDE, PDE3) may interact less strongly with this nitrogen. The PDE4 and PDE3 both interact with cAMP through the 6-amino group, which most likely serves as a hydrogen bond donor. PDE4 and PDE3 appear to be able to bind to the anti-conformer of cAMP, whereas the PDE1 and PDE2 bind the syn-conformer. The CaM-PDE exhibits no appreciable specificity for any of the analogs tested, showing little or no interaction with the 6-amino group or with any of the ring nitrogens. Large differences exist in the nucleotide-binding requirements for the PDE catalytic sites, compared with the regulatory sites of cAMP-dependent protein kinase and the catabolite activator protein.
Butt
E
E
Universität Würzburg, Klinische Forschergruppe, Germany.
Beltman
J
J
Becker
D E
DE
Jensen
G S
GS
Rybalkin
S D
SD
Jastorff
B
B
Beavo
J A
JA
D000818:Animals; D001665:Binding Sites; D002384:Catalysis; D002460:Cell Line; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011506:Proteins / Q000378:metabolism*
D011506:Proteins; D000242:Cyclic AMP; D010727:Phosphoric Diester Hydrolases