ATPase-promoting dead end inhibitors of the cAMP-dependent protein kinase.
The cAMP-dependent protein kinase is a bifunctional enzyme, catalyzing the phosphorylation of the serine and threonine residues in peptides and proteins (kinase activity) as well as the phosphorylation of water (ATPase activity). We have found that several peptides, which serve as inhibitors of the kinase reaction, will either maintain or enhance the ATPase reaction catalyzed by the enzyme. Positively charged dipeptides (e.g. Arg-Arg), as well as small guanidino-containing compounds (e.g. guanethidine) block protein kinase activity yet enhance ATPase activity up to 3.5-fold over that exhibited by the enzyme in the absence of these compounds. In contrast, several nonphosphorylatable peptides, whose primary sequences are based on that of a known substrate (i.e. Leu-Arg-Arg-Ala-Ser-Leu-Gly), such as Leu-Arg-Arg-Ala-Ala-Leu-Gly, Leu-Arg-Arg-Ala-Phe-Leu-Gly, and Leu-Arg-Arg-Ala-Tyr-Leu-Gly, have little or no effect on the rate of the kinase-catalyzed hydrolysis of ATP. An exception to the latter observation is Leu-Arg-Arg-Ala-Cys-Leu-Gly, a cysteine-containing peptide that promotes the protein kinase-catalyzed ATPase reaction by 2.2-fold. We have also found that peptides that possess relatively large amino acid side chain moieties immediately following the arginine dyad (i.e. such as Phe, Tyr, Cys, or Asn at Xaa in Leu-Arg-Arg-Xaa-Ala-Leu-Gly) sharply reduce the rate of enzyme-catalyzed ATP hydrolysis. This suggests that in the presence of peptides containing an -Arg-Arg-Ala- sequence, the enzyme-bound gamma-phosphate of ATP is relatively accessible to water. In contrast, when the latter alanine moiety is replaced by a larger residue, access by water to ATP appears to be hindered. These results indicate that certain structural features associated with the substrate or substrate analog have a profound influence on the manner by which these species interact with the protein kinase. Furthermore, the work described herein demonstrates that it is possible to block the physiologically important kinase reaction and simultaneously promote the energetically wasteful ATPase reaction.
Mendelow
M
M
Department of Chemistry, State University of New York, Buffalo 14214.
Prorok
M
M
Salerno
A
A
Lawrence
D S
DS
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D004151:Dipeptides / Q000494:pharmacology*; D007700:Kinetics; D008433:Mathematics; D008962:Models, Theoretical; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D004151:Dipeptides; D009842:Oligopeptides; D047428:Protein Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D000251:Adenosine Triphosphatases