CHARACTERIZATION OF THE SOLUBLE CYCLIC-NUCLEOTIDE PHOSPHODIESTERASES IN XENOPUS-LAEVIS OOCYTES - EVIDENCE FOR A CALMODULIN-DEPENDENT ENZYME

Cyclic nucleotide phosphodiesterase activities (3'',5''-cyclic-nucleotide 5''-nucleotidohydrolase, EC 3.1.4.17) were found in the 40,000 .times. g supernatant fraction of homogenates of X. laevis oocytes. In the supernatant, the ratio of the specific activity of cAMP phosphodiesterase to that of cGMP phosphodiesterase was 1.1 at the 1 .mu.M substrate level. Two phosphodiesterase forms were isolated by centrifugation on sucrose gradient: a 3-4 S form hydrolyzing specifically cAMP and a 6-7 S form hydrolyzing both cyclic nucleotides (cAMP and cGMP). The activity of the 6-7 S phosphodiesterase was characterized by its activation by 0.1 .mu.M calmodulin purified from beef pancreas in the presence of 50 .mu. M Ca2+. The calmodulin dependence of this form was completely abolished in the presence of 1 mM ethyleneglycol bis(.beta.-aminoethyl ether)-N,N,N'',N''-tetraacetic acid (EGTA). Trifluoperazine at 0.1 mM inhibited both the freshly prepared crude enzyme and the partially purified 607 S form. No effect of cGMP at 3 .mu. was observed on cAMP hydrolysis in the case of the supernatant or that of the partially purified phosphodiesterases. These data show the presence of a calmodulin-dependent phosphodiesterase in the soluble fraction of X. laevis oocytes.