The arginine finger of RasGap helps Gln-61 align the nucleophilic water in GAP-stimulated hydrolysis of GTP

The Ras family of GTPases is a collection of molecular switches that link receptors on the plasma membrane to signaling pathways that regulate cell proliferation and differentiation. The accessory GTPase-activating proteins (GAPs) negatively regulate the cell signaling by increasing the slow intrinsic GTP to GDP hydrolysis rate of Ras. Mutants of Ras are found in 25-30% of human tumors. The most dramatic property of these mutants is their insensitivity to the negative regulatory action of GAPs. All known oncogenic mutants of Ras map to a small subset of amino acids. Gin-61 is particularly important because virtually all mutations of this residue eliminate sensitivity to GAPs. Despite its obvious importance for carcinogenesis, the role of Gin-61 in the GAP-stimulated GTPase activity of Ras has remained a mystery. Our molecular dynamics simulations of the p21 ras-p120GAP-GTP complex suggest that the local structure around the catalytic region can be different from that revealed by the x-ray crystal structure. We find that the carbonyl oxygen on the backbone of the arginine finger supplied in trans by p120GAP (Arg-789) interacts with a water molecule in the active site that is forming a bridge between the NH₂ group of the Gin-61 and the γ-phosphate of GTP. Thus, Arg-789 may play a dual role in generating the nucleophile as well as stabilizing the transition state for P - O bond cleavage.