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

Glass, D. B., Feller, M. J., Levin, L. R., Walsh, D. A. (February 1992) Structural basis for the low affinities of yeast cAMP-dependent and mammalian cGMP-dependent protein kinases for protein kinase inhibitor peptides. Biochemistry, 31 (6). pp. 1728-34. ISSN 0006-2960 (Print)0006-2960 (Linking)

URL: http://www.ncbi.nlm.nih.gov/pubmed/1310617
DOI: 10.1021/bi00121a021

Abstract

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.

Item Type: Paper
Uncontrolled Keywords: Amino Acid Sequence Animals Binding Sites Cattle Cyclic AMP/*pharmacology Cyclic GMP/*pharmacology Kinetics Molecular Sequence Data Myocardium/*enzymology Oligopeptides/metabolism Peptide Fragments/chemistry/*pharmacology Peptides/chemistry/*pharmacology *Protein Kinase Inhibitors Protein Kinases/chemistry/metabolism Saccharomyces cerevisiae/*enzymology Substrate Specificity
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > transcription factor > Cyclic AMP
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes > kinase
organism description > yeast
CSHL Authors:
Communities: CSHL labs
Depositing User: Matt Covey
Date: 18 February 1992
Date Deposited: 01 Oct 2015 19:02
Last Modified: 01 Oct 2015 19:02
Related URLs:
URI: http://repository.cshl.edu/id/eprint/31773

Actions (login required)

Administrator's edit/view item Administrator's edit/view item
CSHL HomeAbout CSHLResearchEducationNews & FeaturesCampus & Public EventsCareersGiving