Unnatural amino acid mutagenesis in eukaryotes

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DESCRIPTION (provided by applicant): The proposed research seeks to expand the power of unnatural amino acid mutagenesis to use in eukaryotic cells. In order to do this, an orthogonal tRNA/synthetase pair will be introduced into a yeast host. tRNA synthetase active site variants will then be screened for novel specificities to interesting unnatural amino acids using a specially designed genetic selection. Once identified, the new pair will be used to deliver unnatural amino acids site specifically in yeast cells. Later, it might be possible to even move successful pairs into mammalian cell lines. The success of this project will open the door to a whole array of exciting structure/function studies of membrane bound receptors and the engineering of proteins with novel chemical or spectroscopic properties.