The catalytic subunit of protein kinase

The type V cyclic GMP phosphodiesterase was partially purified from the high-speed supernatant of guinea-pig lung. The isoenzyme displayed linear kinetics for cyclic GMP hydrolysis, with Km = 2.2 ± 0.2 ,tM and Vmax = 1.2 ± 0.08 nmol/ min per mg. The selective type V phosphodiesterase inhibitor Zaprinast inhibited cyclic GMP hydrolysis with IC50 (concn. giving 50% inhibition) = 0.45 + 0.08 /,M. Isobutylmethylxanthine promoted a 3-fold increase in the binding of cyclic GMP to the isoenzyme. The addition of the catalytic subunit of protein kinase A to an activation cocktail containing the partially purified type V phosphodiesterase resulted in a marked increase in Vmax for cyclic GMP hydrolysis (10-fold at 40 units of protein kinase A). We have suggested that protein kinase A triggers phosphorylation of the phosphodiesterase, which results in activation of phosphodiesterase activity. In addition, the sensitivity to inhibition by Zaprinast is severely decreased (the IC50 for inhibition is 7.5 + 1.1 ,uM), suggesting that the potency of phosphodiesterase inhibitors is effected by phosphorylation of the enzyme.