Structural Basis for Phosphorylation and Lysine Acetylation Crosstalk in a Kinase Motif Associated with Myocardial Ischemia and Cardioprotection [Signal Transduction]

July 9th, 2014 by Parker, B. L., Shepherd, N. E., Trefely, S., Hoffman, N. J., White, M. Y., Engholm-Keller, K., Hambly, B. D., Larsen, M. R., James, D. E., Cordwell, S. J.

Myocardial ischemia and cardioprotection by ischemic pre-conditioning (IPC) induce signal networks aimed at survival, or cell death if the ischemic period is prolonged. These pathways are mediated by protein post-translational modifications (PTM) that are hypothesized to crosstalk with and regulate each other. Phosphopeptides and lysine acetylated peptides were quantified in isolated rat hearts subjected to ischemia or IPC, with and without splitomicin inhibition of lysine deacetylation. We show lysine acetylation (AcetylK)-dependent activation of AMPK, AKT and PKA kinases during ischemia. Phosphorylation and AcetylK sites mapped onto tertiary structures were proximal in >50% of proteins investigated, yet were mutually exclusive in 50 IPC- and / or ischemia-associated peptides containing the KxxS basophilic protein kinase consensus motif. Modifications in this motif were modelled in the C-terminus of muscle-type creatine kinase (CKM). AcetylK increased proximal dephosphorylation by 10-fold. Structural analysis of modified CKM peptide variants by 2D-NMR revealed stabilization via a lysine-phosphate salt bridge, which was disrupted by AcetylK resulting in backbone flexibility and increased phosphatase accesibility.
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