Proteome-wide prediction of PKA phosphorylation sites in eukaryotic kingdom

Protein phosphorylation is one of the most essential post-translational modifications (PTMs), and orchestrates a variety of cellular functions and processes. Besides experimental studies, numerous computational predictors implemented in various algorithms have been developed for phosphorylation sites prediction. However, large-scale predictions of kinase-specific phosphorylation sites have not been successfully pursued and remained to be a great challenge. In this work, we raised a “kiss farewell” model and conducted a high-throughput prediction of cAMP-dependent kinase (PKA) phosphorylation sites. Since a protein kinase (PK) should at least “kiss” its substrates and then run away, we proposed a PKA-binding protein to be a potential PKA substrate if at least one PKA site was predicted. To improve the prediction specificity, we reduced false positive rate (FPR) less than 1% when the cut-off value was set as 4. Successfully, we predicted 1387, 630, 568 and 912 potential PKA sites from 410, 217, 173 and 260 PKA-interacting proteins in Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster and Homo sapiens, respectively. Most of these potential phosphorylation sites remained to be experimentally verified. In addition, we detected two sites in one of PKA regulatory subunits to be conserved in eukaryotes as potentially ancient regulatory signals. Our prediction results provide an excellent resource for delineating PKA-mediated signaling pathways and their system integration underlying cellular dynamics and plasticity.