Research ArticleBiochemistry

Engineering allosteric regulation in protein kinases

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Sci. Signal.  06 Nov 2018:
Vol. 11, Issue 555, eaar3250
DOI: 10.1126/scisignal.aar3250

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Hotspots of kinase regulation

The addition of phosphate moieties can alter protein conformation, function, abundance, and localization, thereby regulating a myriad of cellular behaviors and responses. These sites are often mutated in cancer and other diseases. Pincus et al. mapped the residue alignment and patterning of phosphoregulatory sites in eukaryotic kinases and determined that kinases share a conserved architecture, such that introducing a target motif of the kinase PKA at surface sites within two different yeast kinases altered their activity, pathway interactions, and subcellular localization, as well as the overall response of the yeast to pheromone and osmotic signals, in a PKA-dependent manner. These findings, which reveal insights into kinase evolution, also have implications for biological engineering and medicine.

Abstract

Phosphoregulation, in which the addition of a negatively charged phosphate group modulates protein activity, enables dynamic cellular responses. To understand how new phosphoregulation might be acquired, we mutationally scanned the surface of a prototypical yeast kinase (Kss1) to identify potential regulatory sites. The data revealed a set of spatially distributed “hotspots” that might have coevolved with the active site and preferentially modulated kinase activity. By engineering simple consensus phosphorylation sites at these hotspots, we rewired cell signaling in yeast. Using the same approach with a homolog yeast mitogen-activated protein kinase, Hog1, we introduced new phosphoregulation that modified its localization and signaling dynamics. Beyond revealing potential use in synthetic biology, our findings suggest that the identified hotspots contribute to the diversity of natural allosteric regulatory mechanisms in the eukaryotic kinome and, given that some are mutated in cancers, understanding these hotspots may have clinical relevance to human disease.

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