Activation of protein kinase isoenzymes under near physiological conditions: Evidence that both types (A and B) of cAMP binding sites are involved in the activation of protein kinase by cAMP and 8-N 3-cAMP

Abstract

cAMP-dependent protein kinase I and II (cAKI and cAKII) were incubated under near physiological conditions in the presence of various concentrations of 8-N 3-c[ 3H]AMP or c[ 3H]AMP. Both types (A and B) of cyclic nucleotide binding sites of cAKI or cAKII were occupied to a similar extent and the degree of their occupation correlated with the degree of kinase activation. cAKI cAKII bound cAMP in an apparent positively cooperative manner in the presence of Mg 2+, ATP. 8-N 3-c[ 3H]AMP dissociated several orders of magnitude faster from site A than site B of the regulatory moeity of cAKII, and was photo-incorporated only when bound to site B.