Small compounds and metabolites as conformational modulators of full-length PDK1 (Póster)

LISSY GROSS; MARIANA SACERDOTI; ALEJANDRO E. LEROUX; KEVIN G. HICKS; JARED RUTTER; MELISSA A. GRÄWERT; DMITRI SVERGUN; ABHIJEET GHODE; GANESH S. ANAND; SEBASTIÁN KLINKE; RICARDO M. BIONDI

Evento online debido a la pandemia de coronavirus

Congreso; Reunión conjunta LVI SAIB - XV SAMIGE; 2020

Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB) y Sociedad Argentina de Microbiología General (SAMIGE)

Protein kinases are very important ON-OFF switches in cells. Dysregulation of their activities often lead to diseases, such as cancer or diabetes. Therefore, protein kinases are tightly and selectively regulated. Despite the large knowledge on the role of kinases, the structures of full-length protein kinases are vastly unknown and very few studies have investigated multidomain protein kinases in solution from a structural perspective. Phosphoinositide-dependent protein kinase 1 (PDK1) is a master AGC kinase that phosphorylates at least other 23 AGC kinases, being PKB/Akt the most relevant substrate downstream of PI3-kinase, important for growth and cell survival and a drug target for cancer treatment. PDK1 consists of an N-terminal protein kinase catalytic domain, followed by a linker and a C-terminal PH domain. Over the years, our laboratory studied and characterized in detail the catalytic domain of PDK1. In particular,we employed a chemical biology approach to describe the allosteric communication between a hydrophobic regulatory site, termed PIF-pocket, and the ATP-binding site. We developed small compounds that bind to the PIF-pocket regulatory site and allosterically affect the ATP-binding site. We also described small compounds that bind at the ATP-binding site and allosterically affect interactions at the regulatory site. Our previous studies opened the possibility that metabolites binding at the ATP-binding site could allosterically modulate protein kinase interactions and the formation of protein kinase complexes. This new regulation approach implies that the metabolic state of cells could be linked to the regulation of cell signalling. Here I present the crystal structure of the catalytic domain of PDK1 in complex with a metabolite bound to the ATP-binding site, DMP, and compare the structure of the complex and allosteric effects of this metabolite to the crystal structures and allosteric effects of the metabolites Adenine and Adenosine, which also bind at the ATP-binding site. In addition to this, we identified small compounds that can modulate the conformation of full length PDK1. To study this ligand-dependent conformational modulation, we investigated the effect of these small compounds on full length PDK1 using Hydrogen/Deuterium exchange studies as well as small-angle X-ray scattering (SAXS) experiments. Finally, I will present our preliminary models on full length PDK1.