Diastereomers of adenosine 3′,5′‐monothionophosphate (cAMP[S] antagonize the activation of cGMP‐dependent protein kinase

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Abstract
CGMP-dependent protein kinase contains 4 cGMP-binding sites which are homologeous to the 4 cAMP-binding sites of cAMP-dependent protein kinase. The interaction of the diastereomers of adenosine 3'',5''-thionophosphate, (PS)-cAMP[S] and (PR)-cAMP[S], with cGMP-dependent protein kinase was studied. Autophosphorylation of cGMP-dependent protein kinase is stimulated by cAMP and (PS)-cAMP[S] with apparent KA values of 7 and 94 .mu.M, respectively. cAMP-stimulated autophosphorylation is inhibited competitively by (PR)-cAMP[S] with a Ki value of 15 .mu.M. The phosphorylation of the peptide substrate (Leu-Arg-Arg-Ala-Ser-Leu-Gly) is stimulated by cGMP (.apprx. KA 1 .mu.M) and cAMP (.apprx. KA 98 .mu.M) but neither by the (PR) nor (PS) stereomer of cAMP[S]. (PR)-cAMP[S] and (PS)-cAMP[S] inhibit competitively cAMP- or cGMP-stimulated phosphorylation of the peptide substrate with Ki values of 52 .mu.M and 73 .mu.M, respectively. (PS)-cAMP[S] stimulates the phosphorylation of the peptide substrate by an autophosphorylated enzyme. Binding of [3H]cGMP-dependent protein kinase is inhibited by (PS)-cAMP[S] and (PR)-cAMP[S] with IC50 values of 200 and 15 .mu.M, respectively. Apparently both diastereomers of cAMP[S] bind to cGMP-dependent protein kinase, (PR)-cAMP[S] has properties of a pure antagonist; (PS)-cAMP[S] has properties of a partial agonist. The results provide further evidence that autophosphorylation of the ezyme affects the interaction between the cGMP-binding sites and the catalytic center of the enzyme by facilitating the activation of the phosphotransferase reaction.