Characterization of the Influence of a Small Molecule on a Ras-Related Protein-Protein Interaction

Abstract

Mutations and/or abnormal expression affecting Ras-related protein interactions may influence their roles in cell-signaling activity. Thusly, it remains important to characterize molecular features of Ras protein interactions that dictate their function as ‘timing switches’ in signal transduction pathways. This is a critical step towards targeting the inhibition of cell transformation as well as controlling abnormal signaling activity of tumor-causing Ras variants. Cell division cycle 42 (Cdc42) is a member of the Ras family of proteins. Previous structural studies on a Cdc42 construct, Cdc42(T35A), from this laboratory have outlined differences in conformational dynamics in an important region that interacts with various effectors, known as the Switch 1 region, in this variant, as compared to wild type Cdc42. These differences correlated to a reduced binding of an effector protein that inhibits GTP hydrolysis. As such, the targeting of a small molecule towards an important effector-binding region of Cdc42 could facilitate changes in local conformational dynamics without affecting the overall stability of the Ras protein. Moreover, this effort could provide valuable contributions toward their use as Ras inhibitors. We are currently using biochemical and biophysical approaches to characterize the influence of a small molecule (ZCL278) has on the interaction between Cdc42 and a GTPase inhibitor Protein. Molecular modeling studies have suggested that this small molecule can bind to the Switch 1 region of Cdc42 and “Mimic” an interaction with an effector protein. Our preliminary data outlines the influence this small molecule has on Cdc42's interaction with an effector protein that inhibits Cdc42-stimulated GTPase activity. We also present results that examine ZCL278's influence on a Cdc42 variant with altered dynamics in the Switch 1 region.