Structural basis for the low affinities of yeast cAMP-dependent and mammalian cGMP-dependent protein kinases for protein kinase inhibitor peptides

Biochemistry. 1992 Feb 18;31(6):1728-34. doi: 10.1021/bi00121a021.


Affinities of the catalytic subunit (C1) of Saccharomyces cerevisiae cAMP-dependent protein kinase and of mammalian cGMP-dependent protein kinase were determined for the protein kinase inhibitor (PKI) peptide PKI(6-22)amide and seven analogues. These analogues contained structural alterations in the N-terminal alpha-helix, the C-terminal pseudosubstrate portion, or the central connecting region of the PKI peptide. In all cases, the PKI peptides were appreciably less active as inhibitors of yeast C1 than of mammalian C alpha subunit. Ki values ranged from 5- to 290-fold higher for the yeast enzyme than for its mammalian counterpart. Consistent with these results, yeast C1 exhibited a higher Km for the peptide substrate Kemptide. All of the PKI peptides were even less active against the mammalian cGMP-dependent protein kinase than toward yeast cAMP-dependent protein kinase, and Kemptide was a poorer substrate for the former enzyme. Alignment of amino acid sequences of these homologous protein kinases around residues in the active site of mammalian C alpha subunit known to interact with determinants in the PKI peptide [Knighton, D. R., Zheng, J., Ten Eyck, L. F., Xuong, N-h, Taylor, S. S., & Sowadski, J. M. (1991) Science 253, 414-420] provides a structural basis for the inherently lower affinities of yeast C1 and cGMP-dependent protein kinase for binding peptide inhibitors and substrates. Both yeast cAMP-dependent and mammalian cGMP-dependent protein kinases are missing two of the three acidic residues that interact with arginine-18 in the pseudosubstrate portion of PKI. Further, the cGMP-dependent protein kinase appears to completely lack the hydrophobic/aromatic pocket that recognizes the important phenylalanine-10 residue in the N-terminus of the PKI peptide, and binding of the inhibitor by the yeast protein kinase at this site appears to be partially compromised.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Cattle
  • Cyclic AMP / pharmacology*
  • Cyclic GMP / pharmacology*
  • Kinetics
  • Molecular Sequence Data
  • Myocardium / enzymology*
  • Oligopeptides / metabolism
  • Peptide Fragments / chemistry
  • Peptide Fragments / pharmacology*
  • Peptides / chemistry
  • Peptides / pharmacology*
  • Protein Kinase Inhibitors*
  • Protein Kinases / chemistry
  • Protein Kinases / metabolism
  • Saccharomyces cerevisiae / enzymology*
  • Substrate Specificity


  • Oligopeptides
  • Peptide Fragments
  • Peptides
  • Protein Kinase Inhibitors
  • protein kinase inhibitor peptide
  • kemptide
  • Cyclic AMP
  • Protein Kinases
  • Cyclic GMP