Separation and characterization of a novel isoenzyme of cyclic nucleotide phosphodiesterase from rat cerebrum

Abstract

Anion-exchange chromatography on a Mono-Q column of the supernatant fraction, after ultracentrifugation, from a homogenate of rat cerebrum, prepared under isotonic conditions in the presence of protease inhibitors, yielded a novel isoenzyme of cyclic nucleotide phosphodiesterase (PDE) with properties unlike those of known PDEs. The isoenzyme was insensitive to stimulation by Ca²⁺/calmodulin and cyclic GMP, and it hydrolyzed both cyclic AMP and cyclic GMP with K_M values of 0.109 ± 0.008 μm and 1.78 ± 0.04 μm, respectively. The ratio of V_max of hydrolysis of cyclic GMP to that of cyclic AMP was 1.90 ± 0.07. Nicardipine (PDE I inhibitor), SK&F 94120 (PDE III inhibitor), rolipram (PDE IV inhibitor) and zaprinast (PDE V inhibitor) had very weak inhibitory effects on the PDE activity of the isoenzyme. These results suggest that the isoenzyme is a novel and previously unreported species of PDE, which we tentatively designate PDE VIII.