The goal of the proposal is to establish an in vivo assay to detect membrane protein-protein interaction. This technology is especially important to pharmaceutical industry because most drug targets are membrane proteins. The proposed detection system is based on ornithine decarboxylase (ODC), a key enzyme in the biosynthesis of polyamines. The active form of ODC is a homodimer, but mutations at amino add 316 of mouse ODC prevent the dimer formation. The idea of the proposal is to reassemble the mutant to form a functional dimer by a pair of interacting proteins. The dimer' formation leads to restoring ODC activity which can be used as a marker for selection, such as conferring ODC-deficient cells to grow in a medium with polyamines or rendering wild type ODC cells to grow in the presence of certain amount of ODC inhibitor. Because the distribution of ODC in both cytosol and nucleus, fusion of ODC with membrane proteins can be used for the detection of membrane protein-protein interaction. It will facilitate the discovery of potential drugs that can inhibit the interaction or the search of novel interacted proteins. PROPOSED COMMERCIAL APPLICATION: Since the detection system has no restriction in the requirement of cellular compartment for the system to detect protein-protein interactions, it will be able to detect some of the interactions that are unable to be detected by the yeast two-hybrid system, especially membrane protein-protein interaction. The system can be provided as a commercial kit to the research community for examining protein-protein interaction or searching novel interacted proteins as well as to the pharmaceutical industry for screening new drugs to prevent the protein interaction of clinical significance.
