Methods of identifying peptide agonists or negative antagonists of a G protein coupled receptor (U.S. Patent Number 6,403,305)

The present invention is directed to a strategy to discover small peptides that will activate any G protein coupled receptor (GPCR) or inactivate any constitutively active GPCR. The strategy uses combinatorial peptide libraries to screen millions of random peptides for agonist/negative antagonist activity. The method of the subject invention comprises expressing a peptide of a peptide library tethered to a G protein coupled receptor of interest in a cell, and monitoring the cell to determine whether the peptide is an agonist or negative antagonist of the GPCR of interest. The peptide is tethered to the GPCR by replacing the amino terminus of the GPCR with the amino terminus of a self-activating receptor, and replacing the natural peptide ligand present in the amino terminus of the self-activating receptor with the peptide of the peptide library. In one embodiment for discovery of agonists, a ligand of the self-activating receptor is used to cleave the resulting amino terminus to expose the peptide of the peptide library. In another embodiment for discovery of agonists or negative antagonists, the GPCR construct ends at the peptide so the peptide is always exposed. Preferably, the self-activating receptor is the thrombin receptor and the ligand of the self-activating receptor is thrombin.