Quantitative labeling of the regulatory subunit of type II cAMP-dependent protein kinase from bovine heart by a photoaffinity analog.
Several methods were compared for estimating the amount of regulatory subunit of an 800-fold purified Type II cAMP-dependent protein kinase from bovine heart. These methods included a reversable binding assay using either cAMP, or 8-N3-[32P]cAMP, photoaffinity labeling with 8-N3-[32P]cAMP, and autophosphorylation of the regulatory subunit of the enzyme. Although the regulatory subunit had a slightly lower affinity for 8-N3-cAMP than for cAMP, the total amount of regulatory subunit could be determined by each of the procedures examined. The results indicate that the photoaffinity analog 8-N3-[32P]cAMP is able to label quantitatively all cAMP-binding sites of the regulatory subunit of this cAMP-dependent protein kinase.
D000345:Affinity Labels; D000818:Animals; D001386:Azides; D001665:Binding Sites; D002417:Cattle; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D009206:Myocardium / Q000201:enzymology*; D010766:Phosphorylation; D010777:Photochemistry; D011494:Protein Kinases / Q000378:metabolism*
D000345:Affinity Labels; D001386:Azides; C017042:8-azidoadenosine-3',5'-monophosphate; D000242:Cyclic AMP; D011494:Protein Kinases