Calmodulin-activated cyclic nucleotide phosphodiesterase from brain. Changes in molecular size assessed by gel filtration and electrophoresis.

The calmodulin-activated cyclic nucleotide phosphodiesterase from bovine brain cortex has been extensively purified by two methods, both of which yield preparations containing a single major polypeptide of 58,000 daltons as judged by gel electrophoresis under denaturing conditions. The preparations have similar specific activities in the presence of calmodulin (plus Ca2+) but differ in the degree to which they are activated by this effector. Thus, the method used for purification appeared to influence the relative basal activity of a preparation. On gel permeation chromatography of supernatants immediately after preparation, the phosphodiesterase activity behaved as a protein of -50,000 to 60,000 daltons. After storage of the supernatant for several days at 4 “C, the enzyme had the chromatographic behavior expected for a protein of 120,000-140,000 daltons and was stimulated to a greater degree by Ca2+ plus calmodulin. The apparent half-time for this transition at neutral pH was 24 to 30 h. Storage of a highly purified preparation of phosphodiesterase resulted in a time-dependent decrease in the degree of activation by calmodulin with no change in the ability of the enzyme to be retained on calmodulinSepharose or of the peptide pattern observed on gel electrophoresis under denaturing conditions. During this period, gel electrophoresis of the native protein revealed changes consistent with the transition from a predominantly oligomeric to a monomeric form. All of our observations support the ypothesis that the phosphodiesterase exists as interconvertible monomeric and oligomeric species and that these are activated to different degrees by calmodulin.