Binding proteins for adenosine 3':5'-cyclic monophosphate in bovine adrenal cortex.
Five peaks of cyclic AMP-binding activity could be resolved by DEAE-cellulose chromatography of bovine adrenal-cortex cytosol. Two of the binding peaks co-chromatographed with the catalytic activities of cyclic AMP-dependent protein kinases (ATP-protein phosphotransferase, EC 2.7.1.37) of type I or type II respectively. A third binding protein was eluted between the two kinases, and appeared to be the free regulatory moiety of protein kinase I. Two of the binding proteins for cyclic AMP, sedimenting at 9S in sucrose gradients, could also bind adenosine. They bound cyclic AMP with an apparent equilibrium dissociation constant (K(d)) of about 0.1mum, and showed an increased binding capacity for cyclic AMP after preincubation in the presence of K(+), Mg(2+) and ATP. The two binding proteins differed in their apparent affinities for adenosine. The isolated regulatory moiety of protein kinase I had a very high affinity for cyclic AMP (K(d)<0.1nm). At low ionic strength or in the presence of MgATP, the high-affinity binding of cyclic AMP to the regulatory subunit of protein kinase I was decreased by the catalytic subunit. At high ionic strength and in the absence of MgATP the high-affinity binding to the regulatory subunit was not affected by the presence of catalytic subunit. Under all experimental conditions tested, dissociation of protein kinase I was accompanied by an increased affinity for cyclic AMP. To gain some insight into the mechanism by which cyclic AMP activates protein kinase, the interaction between basic proteins, salt and the cyclic nucleotide in activating the kinase was studied.
D000241:Adenosine / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000302:Adrenal Cortex / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002848:Chromatography, DEAE-Cellulose; D000242:Cyclic AMP / Q000378:metabolism; D004789:Enzyme Activation; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D009994:Osmolar Concentration; D011494:Protein Kinases / Q000378:metabolism; D011953:Receptors, Cyclic AMP / Q000378:metabolism*
D011953:Receptors, Cyclic AMP; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D008274:Magnesium; D000241:Adenosine