Effects of fatty acids on activity of cGMP-stimulated cyclic nucleotide phosphodiesterase from calf liver

Abstract

Effects of fatty acids, prostaglandins, and phospholipids on the activity of purified cGMP-stimulated cyclic nucleotide phosphodiesterase from calf liver were investigated. Prostaglandins A 2, E 1, E 2, F 1α, and F 2α, thromboxane B 2, and most phospholipids were without effect; lysophosphatidylcholine was a potent inhibitor. Several saturated fatty acids (carbon chain length 14–24), at concentrations up to 1 m m, had little or no effect on hydrolysis of 0.5 μ m [ 3H]cGMP or 0.5 μ m [ 3H]cAMP with or without 1 μ m cGMP. In general, unsaturated fatty acids were inhibitory, except for myristoleic and palmitoleic acids which increased hydrolysis of 0.5 μ m [ 3H]cAMP. The extent of inhibition by cis-isomers correlated with the number of double bonds. Increasing concentrations of palmitoleic acid from 10 to 100 μ m increased hydrolysis of [ 3H]cAMP with maximal activation (60%) at 100 μ m; higher concentrations were inhibitory. Palmitoleic acid inhibited cGMP hydrolysis and cGMP-stimulated cAMP hydrolysis with IC 50 values of 110 and 75 μ m, respectively. Inhibitory effects of palmitoleic acid were completely or partially prevented by equimolar α-tocopherol. Palmitelaidic acid, the trans isomer, had only slightly inhibitory effects. The effects of palmitoleic acid (100 μ m) were dependent on substrate concentration. Activation was maximal with 1 μ m [ 3H]cAMP and was reduced with increasing substrate; with >10 μ m cAMP, palmitoleic had no effect. Inhibition of cGMP hydrolysis was maximal at 2.5 μ m cGMP and was reduced with increasing cGMP; at >100 μ m cGMP palmitoleic acid increased hydrolysis slightly. Palmitoleic acid did not affect apparent K m or V max for cAMP hydrolysis, but increased the apparent K m (from 17 to 60 μ m) and V max for cGMP hydrolysis with little or no effect on the Hill coefficient for either substrate. These results suggest that certain hydrophobic domains play an important role in modifying the catalytic specificity of the cGMP-stimulated phosphodiesterase for cAMP and cGMP.