REGULATION OF PDK1 KINASE BY SMALL MOLECULES AND METABOLITES: BIDIRECTIONAL ALLOSTERY AND BEYOND

SACERDOTI, M; GROSS, LZF; GHODE, A; HICKS, KG; RUTTER, J; ANAND, GS; LEROUX, AE; BIONDI, RM

Conferencia; Translational Chemical Biology; 2020

Nature Conferences and Dana-Farber Cancer Institute

Phosphoinositide-dependentprotein kinase 1 (PDK1) is a master kinase of the PI3-kinase signalling pathwaythat phosphorylates at least 23 other evolutionary related AGC kinases. Phosphorylationby PDK1 is required for the activity of all substrates: they are phosphorylatedeither constitutively or with different timing upon PI3-kinase activation. Themechanism of activation of PDK1, mediated by the PIF-pocket regulatory site, is conserved within the large group ofAGC kinases, including the isoforms of PKC, Akt, SGK, S6K, RSK, MSK, etc.  Over the years, our laboratory has used achemical and structural biology approach to study and characterize thebidirectional allosteric regulation between the PIF-pocket and the ATP-Binding site on the catalytic domain ofPDK1. Compounds that we have developed to bind at the PIF-pocket activate or inhibit AGC kinases by allostericallyaffecting the ATP-Binding site. Furthermore, small compounds, i.e. PS653, anddrugs developed by the pharmaceutical industry to bind at the ATP-Binding sitecan allosterically enhance or inhibit interactions at the PIF-pocket. Indeed, the same bidirectional allosteric mechanism ofregulation that drives conformational changes from a regulatory site to theATP-Binding site can be exploited by drugs targeting the latter to disruptprotein kinase interactions. We next employed biochemical and structuraltechniques, in silico moleculardynamics and hydrogen/deuterium exchange (HDX) approaches to investigatewhether metabolites could also bind at the ATP-Binding site and allostericallymodulate protein kinase interactions. We found that Adenosine, which binds at theATP-Binding site, allosterically enhances interactions at the PIF-pocket. We demonstrate thatAdenosine and PIFtide (a peptidederived from the hydrophobic motif of a PDK1 substrate that binds to the PIF-pocket) synergistically enhance theinteraction of one another. Finally, we investigate the effect of distinct metabolites,which bind at the ATP-binding site, on properties of full length PDK1. Our in vitro studies suggest that theallosteric mechanism of regulation could possibly sense the cellular metabolicstate and alter the dynamic formation of protein kinase complexes,physiologically modulating cellular signaling by a novel mechanism.