Rationale:  Many kinases phosphorylate multiple substrates. To assess the role of an individual phosphorylation event without using laborious mutagenesis studies, selective pharmacological tools that inhibit phosphorylation of specific one substrate are needed. We focused on delta protein kinase C (dPKC) because dPKC activation after heart attack leads to cardiac damage. We reasoned that since pyruvate dehydrogenase kinase (PDK) phosphorylation by dPKC leads to inhibition of ATP production, it might be critical in the injurious effect dPKC after heart attack. To determine if of the many substrates of dPKC, PDK is the substrate that mediates this injurious effect in the heart, we designed a selective inhibitor of dPKC binding and phosphorylation of PDK.Methods : We designed a short peptide inhibitor that mimics a unique interaction between PDK and dPKC to selectively inhibit dPKC phosphorylation of PDK. This protein-protein interaction inhibitory peptide (PPIIP) was used in vitro, in cells and in intact heart, in models of heart attack.Results : PDK/ dPKC interaction was inhibited by PPIIP in vitro and in vivo. PDK/ dPKC PPIIP selectively inhibited phosphorylation of PDK without affecting phosphorylation of several other dPKC substrates. Treatment with PDK/ dPKC PPIIP inhibited myocardial infarction-induced PDK phosphorylation without inhibiting the phosphorylation of other dPKC substrates. Further, PDK/ dPKC PPIIP treatment led to a 50% reduction in infarct size, in release of cardiac enzyme (a measure of cell lysis) and in JNK phosphorylation, all markers of cardiac injury.Conclusion : PDK/ dPKC PPIIP is a selective inhibitor of PDK phosphorylation by dPKC. Of the several dPKC-mediated phosphorylation events following heart attack, dPKC-induced PDK phosphorylation is sufficient to cause cardiac injury as a selective inhibitor of this phosphorylation event alone was sufficient to provide great reduction in dPKC-mediated cardiac injury after heart attack.

Inhibiting one substrate at a time: Bio-engineering of d protein kinase C to modulate specific interactions and phosphorylations

248th National Meeting (2014)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.