Tryptic peptide mapping studies on the regulatory subunits of type II protein kinases from cerebral cortex and heart. Evidence for overall structural divergence and differences in the autophosphorylation and cAMP-binding domains.
Regulatory subunits of type II cAMP-dependent protein kinases (RII) (EC 2.7.1.37) from bovine brain and heart exhibit similar physicochemical and functional properties in vitro. However, the two forms of RII are markedly different in their (a) antigenic determinants, (b) cell and tissue distribution, and (c) subcellular localization. This suggests that each of these cAMP-binding proteins may possess some unique structural features. To assess the degree of overall divergence between the primary structures of brain RII and heart RII, tryptic peptides derived from the two proteins were mapped by reverse phase HPLC on a C18 column. When the column effluent was monitored at 280 nm, 15 peptides were found only in the heart RII digest, while 5 other peptides were obtained only from brain RII. More complex HPLC profiles were observed by following peptide absorbance at 210 nm, but a similar level of diversity was apparent: 13 brain-RII-specific and 15 heart-RII-specific tryptic peptides were identified and resolved with a gradient (0-50%) of acetonitrile in 0.1% trifluoroacetic acid. In complementary experiments, classical two-dimensional mapping analyses revealed that several 32P-labeled tryptic fragments derived from autophosphorylated and photoaffinity-labeled brain RII were separate and distinct from the 32P-peptides isolated from similarly treated heart RII. The HPLC mapping data document a structural basis for the immunological disparity between brain RII and heart RII and suggest that the two cAMP-binding proteins are different proteins rather than interconvertible forms of a single protein.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D002540:Cerebral Cortex / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D000242:Cyclic AMP / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D009206:Myocardium / Q000201:enzymology*; D010446:Peptide Fragments / Q000032:analysis; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D014357:Trypsin
D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D010446:Peptide Fragments; C015482:protein kinase modulator; D000242:Cyclic AMP; D011494:Protein Kinases; D014357:Trypsin