Chemical complementation: a genetic selection system in yeast for drug discovery, protein and enzyme engineering

Abstract

Drug discovery and protein engineering would both benefit from a general method of applying the power of genetic selection. To this end, we have developed chemical complementation. This yeast-based three-component system (a derivative of the yeast two-hybrid system) links the survival of cells to the presence of a specific small molecule. This system comprises a human nuclear receptor protein, its coactivator protein, and a small molecule ligand, where the nuclear receptor and coactivator associate and activate transcription only in the presence of the small molecule ligand. This chemical complementation system is highly sensitive (producing growth at 10 nM ligand concentration), has zero background (no growth without ligand) and high dynamic range (growth density and time equivalent to Gal4 induced growth). This system is ideal for drug discovery and protein engineering. For drug discovery, positive and negative chemical complementation assays are described for discovering nuclear receptor agonists and antagonists. Nuclear receptors are currently targets for ~10% of commonly prescribed drugs and potentially many more. Drug discovery assays may be extended to enzyme targets by engineering receptors that activate transcription in response to the small molecule product of the enzyme-catalyzed reaction. For protein engineering, chemical complementation offers a general method of engineering receptors that activate transcription in response to arbitrary small molecules. Engineered proteins will have broad uses as biosensors, in metabolic engineering, and gene therapy.