Differential binding of the regulatory subunits (RII) of cAMP-dependent protein kinase II from bovine brain and muscle to RII-binding proteins.
The association of regulatory subunits (RII) of Type II cAMP-dependent protein kinase from bovine cerebral cortex (RII-B) and bovine cardiac and skeletal muscle (RII-H) with specific binding proteins in bovine brain cytosol and purified brain microtubules was demonstrated using a solid phase binding assay. RII-binding proteins present in bovine cerebral cortex were immobilized on nitrocellulose filters after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incubation of the filters with 32P-labeled regulatory subunits showed that both RII-B and RII-H interact with the 75,000-dalton calmodulin-binding protein (P75) and microtubule-associated protein 2 (MAP-2). However, significant differences in binding affinities and capacities were observed. RII-B displayed a higher affinity for P75 compared to RII-H while RII-H preferentially bound to MAP-2. Quantitation of radioactive RII bound to MAP-2 showed that MAP-2 bound 4-6 times more RII-H than RII-B. The differential binding affinities and capacities of RII-H and RII-B for MAP-2 were not affected by autophosphorylation since both phospho and dephospho forms of RII displayed the same binding characteristics. Competitive binding studies suggest that RII-H and RII-B bind to the same sites on MAP-2. The biochemical basis for the differential binding of RII-B and RII-H to the same sites of MAP-2 is unknown. However, other high affinity RII-binding proteins present in cerebral cortex (i.e. P75) might affect the affinity of RII-B for MAP-2. 32P-RI did not bind to P75 nor MAP-2 under the conditions used.
D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000906:Antibodies / Q000378:metabolism; D001667:Binding, Competitive; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D002417:Cattle; D002540:Cerebral Cortex / Q000201:enzymology*; D003600:Cytosol / Q000201:enzymology; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000201:enzymology; D009132:Muscles / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology; D009419:Nerve Tissue Proteins; D011494:Protein Kinases / Q000276:immunology / Q000378:metabolism*
D054758:A Kinase Anchor Proteins; C484064:AKAP5 protein, Bos taurus; D048868:Adaptor Proteins, Signal Transducing; D000906:Antibodies; D002148:Calmodulin-Binding Proteins; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; D011494:Protein Kinases