The exchange rate of unlabeled adenosine 3', 5'-monophosphate (cAMP) with labeled [3H]cAMP in the dimeric regulatory subunit-cAMP complex of cAMP-dependent protein kinase, type I, purified from rabbit skeletal muscle is described by using the equilibrium isotope exchange technique. Results indicate that the rate of exchange carried out in the absence of the catalytic subunit (C) is rather slow with a half-life of ~870 s. This slow exchange rate is not affected by the presence of MgATP (50 µM). However, when both MgATP (50 µM) and C (1-13 nM) are present, the rate of isotope exchange is observed to increase markedly. Furthermore, less than stoichiometric amounts of C are required for the increase in the rate of CAMP exchange, indicating that the effect of C on the rate enhancement is a catalytic process. These results indicate that in the presence of MgATP, a ternary complex between C and regulatory subunit-cAMP complex must be formed, and a dynamic equilibrium between the ternary complex and its dissociable species must be reached within seconds. On the basis of our kinetic data, it is proposed that the formation of this ternary complex intermediate allows the rapid activation or the inactivation of cAMP-dependent protein kinase following changes in the cellular CAMP levels.
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