Daniel Calero, University of Pittsburgh
Abstract
Time Resolved Experiments of RAS oncogenic mutants
Daniel Calero1, Paola Zinser1, Guowu Lin1, Silvia Russi2, Aina Cohen2 and Guillermo Calero1
1 Department of Structural Biology, University of Pittsburgh School of Medicine, 2Stanford Synchrotron Radiation Lightsource.
The RAS protein is a monomeric small GTPase involved in cellular proliferation. It has been shown that RAS is frequently mutated in lung, colon, and pancreatic cancer; this makes RAS the most commonly mutated protein in human cancers. The RAS protein can exist in two forms: The GTP-bound RAS (active form), or the GDP-bound RAS (inactive form). RAS can break down GTP (through a hydrolysis process) to form GDP; however, RAS dissociates GDP very slowly. RAS activation is facilitated by the family of guanine nucleotide exchange factors (GEFs). In particular the GEF protein SOS, a downstream effector of the growth-factor activated receptor tyrosine kinases, activates KRAS (Kirsten rat sarcoma virus). Upon activation, KRAS will trigger downstream signaling events through the RAF-MEK-ERK pathway leading to cell
proliferation. In this project, I have used time time-resolved X-ray crystallography to study the intrinsic hydrolytic activity of KRAS and its oncogenic G12C mutant to capture its active site residues during the process of GTP hydrolysis. Understanding the mechanisms of nucleotide hydrolysis by KRAS in real-time will give critical information that could help develop novel drugs to inhibit the deadliest cancer protein.