Abstract
Over the last 20 years, protein kinases have been at the forefront of drug discovery programs in the pharmaceutical industry. Almost all protein kinase inhibitor programs were originally directed to the ATP‐binding site, while more recently allosteric drug developments show improved selectivity. Over the years we investigated the molecular mechanisms of regulation of a large group of kinases, termed AGC kinases (PDK1, PRKs, aPKCs, S6K, SGK, Akt/PKB, RSK, etc.). We identified a regulatory site in the small lobe of PDK1, termed “PIF‐binding pocket” that participates in the docking interaction of PDK1 with a subset of substrates, i.e. S6K, SGK, but not PKB/Akt. In addition, the PIF‐pocket of PDK1 ‐and the equivalent PIF‐pocket site in other AGC kinases‐ participates in the mechanism of activation and inhibition of these kinases, by phosphorylation or interaction with other domains. The binding of synthetic small compounds to the PIF‐pocket can “close” the kinase domain and allosterically “activate” PDK1 in vitro, or allosterically affect the ATP‐binding site and be allosteric inhibitors of other AGC kinases. The present research deals with the studies that describe how the allosteric process, from the regulatory site, to the ATP‐binding site can be “reversed” by small compounds binding to the ATP‐binding site. Thus, different compounds binding with high affinity to the ATP‐binding site can produce different “reverse” allosteric effects on the PIF‐pocket of PDK1, i.e. displace or enhance docking interactions, and ultimately can produce different effects in cell signaling. We now extended the study of the allosterc processes in protein kinases outside of the AGC group of protein kinases. Aurora kinase requires interaction with TPX2 for regulation and localization. We found that binding of VX680 at the ATP‐binding site does not affect interaction with TPX2, while the binding of MLN8237 readily displaces TPX2. Thus, besides inhibition of Aurora catalytic activity, MLN8237 also affects the formation of TPX2/mediated Aurora complexes. PLK1 is autoinhibited by an intramolecular interaction between the Polo‐Box Domain and the catalytic domain. Binding of phosphopeptides to the Polo‐Box Domain displaces the intramolecular interaction and enables full enzymatic activity of the kinase domain in vitro. In addition, the interactions mediated by the Polo‐Box Domain provide localization to the kinase. Upon screening of a library of small molecules, we identified 1‐ small compounds that bind to the ATP‐binding site and enhance the interaction of the Polo‐box domain with phosphopeptides, and small compounds that inhibit the interaction of PLK1 with phosphopeptides, thereby stabilizing the closed inactive conformation and inhibiting protein‐protein interactions. We show that allosteric compounds acting on PLK1 and affecting Polo‐Box Domain interactions are specific inhibitors of PLK1 cellular activity independently of their capacity to inhibit PLK1 catalytic activity. The results highlight the widespread opportunity to modulate kinase cellular activity by exploiting bi‐directional allosteric processes. The old allostery concept can still be exploited for innovative rational drug developments to protein kinases.Support or Funding InformationDFG BI1044 12/1; DFG BI1044 13/1; DKTKThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
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