Eukaryotic incorporation of photoreactive amino acids

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DESCRIPTION (provided by applicant): Unnatural amino acids will be incorporated in vivo at specific positions within G-protein coupled receptors in order to fluorescently label proteins and detect and characterize protein/protein and protein/ligand interactions. Specific labeling of a single amino acid within a protein with a fluorescent label will provide information about the local microenvironment of that residue. Labeling with a photocrosslinking amino acid, followed by shotgun proteomics by the Biological Mass Spectrometry Lab at TSRI, will identify ligand interactions of that residue in human orphan receptors expressed in yeast. Photocrosslinking using an unnatural amino acid incorporated in C. elegans will be used to characterize a pathway involved with aging. Work with C. elegans will be in collaboration with Andrew Dillin at The Salk Institute.