Differential and common recognition of the catalytic sites of the cGMP-dependent and cAMP-dependent protein kinases by inhibitory peptides derived from the heat-stable inhibitor protein.
Synthetic peptides corresponding to the active domain of the heat-stable inhibitor protein of cAMP-dependent protein kinase (Cheng, H.-C., Kemp, B. E., Pearson, R. B., Smith, A. J., Misconi, L., Van Patten, S. M., and Walsh, D. A. (1986) J. Biol. Chem. 261, 989-992) were tested as inhibitors of cGMP-dependent protein kinase. The peptides themselves were not substrates. cGMP-dependent protein kinase activity was assayed using histone H2B and two synthetic peptide substrates. Consistent with previous observations of other peptide inhibitors of this enzyme (Glass, D. B. (1983) Biochem. J. 213, 159-164), the inhibitory peptides had no effect on the phosphorylation of histone H2B, but they competitively inhibited cGMP-dependent phosphorylation of the two peptide substrates. The parent inhibitor peptide, PKI(5-24)amide, and a series of analogs had Ki (or IC50) values for cGMP-dependent protein kinase in the range of 15-190 microM. In contrast to their effects on the cAMP-dependent protein kinase, the inhibitory peptides were substantially less potent with cGMP-dependent protein kinase, and potency was reduced by the presence of the NH2-terminal residues (residues 5-13). We conclude that the two protein kinases share a recognition of the basic amino acid cluster within the pseudosubstrate region of the peptide, but that the cGMP-dependent protein kinase does not recognize additional NH2-terminal determinants that make the inhibitor protein extremely potent toward the cAMP-dependent enzyme. Even- when tested at high concentrations and with peptide substrates, the native inhibitor protein did not inhibit cGMP-dependent protein kinase under assay conditions in which the peptides derived from it were inhibitory. Thus, the native inhibitor protein appears to have structural features which block interaction with the cGMP-dependent enzyme and enhance its selectivity for cAMP-dependent protein kinase.
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000494:pharmacology*; D002417:Cattle; D000242:Cyclic AMP / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D006358:Hot Temperature; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D010446:Peptide Fragments / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*
D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D010446:Peptide Fragments; C015482:protein kinase modulator; D000242:Cyclic AMP; D011494:Protein Kinases; D006152:Cyclic GMP