[Cyclic AMP-specific nucleotide phosphodiesterase from the insoluble fraction of the human brain]

The cAMP-specific cyclic nucleotide phosphodiesterase (PDE) from pellet fraction of human brain has been identified. Two alternative methods for isolation and purification of the enzyme are presented. The first method consists of three chromatographic steps (separation on the DEAE-Toyopearl column followed by gel filtrations on AcA-34 and G3000 SW columns) and results in purification of several forms of PDE. One of those is weakly inhibited by cGMP and is insensitive to Ca(2+)-calmodulin or cyclic nucleotides. The second procedure consisting of the chromatographic separation on blue agarose and DEAE-TSK-5PW ion-exchange resin leads to an isolation of several peaks of cAMP-specific PDE. Both methods produce different cAMP-specific forms of PDE with molecular masses ranging from 36 to 47 kDa. These forms differ in kinetic properties (Km = 2.0-7.3 microM, Ki = 1480-5300 microM for cGMP). However, the KicGMP/KmcAMP ratio remains constant for all the peaks of the cAMP-specific enzyme.