INTERACTION OF A NOVEL GDP EXCHANGE INHIBITOR WITH THE RAS PROTEIN

Mutated, tumorigenic Ras is present in a variety of human tumors. Comp ounds that inhibit tumorigenic Ras function may be useful in the treat ment of Ras-related tumors. The interaction of a novel GDP exchange in hibitor (SCH-54292) with the Ras-GDP protein was studied by NMR spectr oscopy. The binding of the inhibitor to the Ras protein was enhanced a t low Mg2+ concentrations, which enabled the preparation of a stable c omplex for NMR study. To understand the enhanced inhibitor binding and the increased GDP dissociation rates of the Ras protein, the conforma tional changes of the Ras protein at low Mg2+ concentrations was inves tigated using two-dimensional H-1-N-15 HSQC experiments. The Ras prote in existed in two conformations in slow exchange on the NMR time scale under such conditions. The conformational changes mainly occurred in the GDP binding pocket, in the switch I and the switch II regions, and were reversible. The Ras protein resumed its regular conformation aft er an excess amount of Mg2+ was added. A model of the inhibitor in com plex with the Ras-GDP protein was derived from intra- and intermolecul ar NOE distance constraints, and revealed that the inhibitor bound to the critical switch II region of the Ras protein.