Selective inhibition of two soluble adenosine cyclic 3',5'-phosphate phosphodiesterases partially purified from calf liver.
"Low Km" cAMP phosphodiesterase and cGMP-stimulated cyclic nucleotide phosphodiesterase activities were partially purified from calf liver supernatant by chromatography on DEAE-cellulose and DEAE-Sepharose and ammonium sulfate precipitation. The low Km phosphodiesterase was not retained on N6-H2N(CH2)2-cAMP-agarose and could be separated from the cGMP-stimulated phosphodiesterase which was absorbed by this matrix. From the proteins that did not bind, two distinct low Km cAMP phosphodiesterases were separated on Ultrogel AcA 34. One form (fraction C) hydrolyzed cAMP with an apparent Km of approximately 0.5 microM and was very sensitive to inhibition by cGMP. Lineweaver-Burk plots of cAMP hydrolysis by a second form (fraction B) were nonlinear, with an apparent low Km component of approximately 2 microM. This form was rather insensitive to inhibition by cGMP. With both fractions, hydrolysis of cAMP relative to cGMP was much greater at low (approximately 1 microM) than at high (approximately 100 microM) substrate concentrations. Maximal velocities for cAMP and cGMP were similar. From sedimentation equilibrium, the apparent weight-average molecular weight of fraction B was estimated as 174000, and that of fraction C was 85000. Another fraction (A) of cAMP phosphodiesterase eluted at the void volume of the AcA 34 column. On the basis of the relative affinities for cAMP and cGMP and inhibition by cGMP, fraction A is most likely an aggregated form of fraction B. No apparent interconversion of fractions A, B, or C was observed on high-performance liquid chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D012368:4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D006152:Cyclic GMP / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D011804:Quinolines / Q000494:pharmacology; D015363:Quinolones; D012995:Solubility
D011804:Quinolines; D015363:Quinolones; D012368:4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone; C021294:cilostamide; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D006152:Cyclic GMP