Phosphoryl Transfer Transition State Computationally Modeled by Mgf3(-) in the Oncogenetically Indicated Gtpase Protein Rhoa and it's Activating Protein Rhoa.Gap

Abstract

GTPase enzymes, which hydrolyze guanosine triphosphate (GTP) to guanosine diphosphate (GDP) and inorganic phosphate (Pi), are involved in a large number of critical cellular processes including proliferation. GTPase Activating Protein (GAP) is responsible for the regulation of GTPase. GTPase proteins, when poorly regulated, can signal for uncontrolled cellular growth and are indicated in oncogenesis [1]. I present a computational model of the GTPase protein RhoA in solution and bound to the regulating protein RhoA.GAP. I have modeled a transition state analog of the GTP to GDP phosphoryl transfer reaction based on the crystal structure of the RhoA:RhoGAP protein complex with GDP and MgF3- [2]. Hybrid QM/MM Car-Parrinello MD simulations were performed on the protein complex with the transition state analog, all in explicit water. The reaction was modeled using thermodynamic integration by increasing the terminal phosphate-oxygen bond. These simulations provide evidence for a dissociative transition state and suggest that MgF3- is the best adduct to date to model the transition states of these types of enzymatic phosphoryl transfer reactions. This model could be further exploited to design transition state mimics for many families of GTPase proteins. Finally, this model has shown some structural effects of binding the RhoA.GAP protein to RhoA. These preliminary results indicate that we can understand better the RhoGAP:RhoA protein interface using computational methods. This model could be further exploited to identify ways to dissociate the errant GTPase:GAP complex by targeting the GTP binding site or even the protein-protein interface itself.