Over the last decade, site-specific incorporation of non-canonical amino acids (ncAAs) into proteins in living cells has emerged as an enabling technology for biological research. Our group focuses on further development and application of this technology to probe and engineer biological processes associated with mammalian cells. Two particularly exciting problems that we are currently applying this technology to are: 1) to probe and engineer the molecular processes associated with the entry of adeno-associated virus into human cells, and 2) to elucidate the roles of various post-translational modification of mammalian proteins. To facilitate these applications, we have: 1) developed unique platforms that enable site-specific incorporation of previously inaccessible ncAAs into proteins expressed in both eukaryotes and E. coli, 2) generated a baculovirus-based vector to efficiently deliver the associated genetic machinery into a wide variety of cells and tissues, 3) performed systematic analysis to identify what limits the overall efficiency of ncAA incorporation in mammalian cells, and 4) created a unique mammalian cell based directed evolution platform to improve the performance of these limiting components.
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