Plasma Membrane Cyclic Adenosine 3′:5′-Monophosphate Phosphodiesterase of Cultured Cells and Its Modification after Trypsin Treatment of Intact Cells

Treatment of intact chicken embryo fibroblasts in tissue culture with trypsin leads to an alteration of the kinetic properties of a particulate cyclic adenosine 3':5'-monophosphate (cyclic AMP) phosphodiesterase. Distribution studies show that this phosphodiesterase is principally located in the plasma membrane. Little activity is found in nuclear, mitochondrial, and microsomal fractions. The trypsin-sensitive phosphodiesterase activity does not function on the exterior of the cell membrane. The Km and Vmax of the enzyme are altered by trypsin treatment of intact cells and this effect is quantitatively reproduced by trypsin treatment of the isolated plasma membrane fraction. The supernatant fraction of chicken embryo fibroblasts contains phosphodiesterases capable of hydrolyzing both cyclic AMP and cyclic GMP. DEAE-cellulose chromatography of the supernatant fraction resolves phosphodiesterase activity into two peaks. One fraction is specific for cyclic AMP and is kinetically similar to the plasma membrane enzyme, whereas the other fraction hydrolyzes both cyclic nucleotides in a Michaelis-Menten fashion. The cyclic AMP-specific phosphodiesterase appears to be under negative cooperative regulation. These studies show that chicken embryo fibroblasts contain a cyclic AMP phosphodiesterase in the plasma membrane, the kinetic properties of which can be altered by an enzyme that acts on the outer cell membrane.