CYCLIC AMP BINDING PROTEINS AND RAS FUNCTION

TWO CAMP BINDING PROTEINS (50K AND 10K DALTONS) THAT ALTER BIOCHEMICAL PROPERTIES OF PROTOON- COGENE RAS IN A CAMP DEPENDENT MANNER WERE ISOLATED FROM BOVINE BRAIN. IN THE PRESENCE OF CAMP, THE 50K DALTON PROTEIN (P50) INHIBITED THE NUCLEOTIDE EXCHANGE OF RAS, WHEREAS THE 10K PROTEIN (P10) INHIBITED NUCLEOTIDE BINDING. IN THE ABSENCE OF CAMP, THESE TWO PROTEINS ARE TOTALLY INACTIVE. IT IS WELL ESTABLISHED THAT PROTOONCOGENE RAS PLAYS AN IMPORTANT ROLE IN CONTROLLING CELL PROLIFERATION AND DIFFERENTIATION. THE INTERACTION BETWEEN THESE TWO PROTEINSAND RAS MAY REPRESENT A NOVEL MECHANISM BY WHICH CELL PROLIFERATION AND DIFFERENTIATION. THE INTERACTION BETWEEN THESE TWO PROTEINS AND RAS MAY REPRESENT A NOVEL MECHANISM BY WHICH CELL PROLIFERATION AND DIFFERENTIATION ARE REGULATED. THE OBJECTIVE OF THIS PHASE I PROJECT IS TO PARTIALLY PURIFY THESE TWO PROTEINS, ESTABLISHED A STANDARD ASSAY AND SCREEN FOR THE COMPOUND OR NATURAL PRODUCTS THAT ARE ABLE TO ALTER THE BIOCHEMICAL AND BIOLOGICAL ACTIVITY OF THESE TWO PROTEINS. THE COMPOUNDS IDENTIFIED IN THIS PHASE I STUDY MAY DEVELOP INTO POTENTIAL ANTI- PROLIFERATIVE DRUGS. IN THE PHASE II PORTION OF THIS GRANT, THESE TWO PROTEINS WILL BE PURIFIED, SEQUENCED AND CLONED. THE BIOLOGICAL SIGNIFICANCE OF THESE TWO PROTEINS WILL BE ELUCIDATED BY INTRODUCTING THESE TWO PROTEINS INTO CULTURED CELLS BY PROTEIN MICROINJECTION AND/OR GENE TRANSFER TECHNIQUES. ALTERNATIVELY, ANTIBODIES THAT ARE ABLE TO NEUTRALIZE THESE TWO PROTEINS WILL BE MICROINJECTED INTO CULTURED CELLS TO DETERMINE THE BIOLOGICAL FUNCTION OF THESE TWO PROTEINS. THE CELL MODEL WILL BE ESTABLISHED TO SCREEN THE COMPOUNDS THAT ARE ABLE TO ALTER THE BIOLOGICAL ACTIVITY OF THESE TWO PROTEINS.