Separation and characterization of a novel isoenzyme of cyclic nucleotide phosphodiesterase from rat cerebrum.
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 KM values of 0.109 +/- 0.008 microM and 1.78 +/- 0.04 microM, respectively. The ratio of Vmax 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.
Mukai
J
J
Department of Molecular and Cellular Pharmacology, Mie University School of Medicine, Japan.
Asai
T
T
Naka
M
M
Tanaka
T
T
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D000818:Animals; D001921:Brain / Q000187:drug effects / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000494:pharmacology; D002852:Chromatography, Ion Exchange; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D051381:Rats
D002147:Calmodulin; D007527:Isoenzymes; D000242:Cyclic AMP; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D006152:Cyclic GMP; D002118:Calcium