Expanding the Genetic Code
Abstract
We are exploring the application of unnatural base pairs (UBPs) as an expansion of the genetic code. To prevent loss of the unnatural base pair during reproduction we will utilize several systems including CRISPR/Cas9. The extended genetic code allows the ribosomal incorporation of several non-canonical amino acids (ncAAs) into peptides. With their broad chemical and functional diversity ncAAs provide a variety of promising applications including protein labeling, photocaging, structure analysis and specific protein interactions. Therefore, the development of a new toolkit for the translational incorporation of ncAAs in E. coli would be a great contribu-tion to the iGEM community. The site-specific incorporation of ncAAs requires the directed evolution of tRNA/aminoacyl-tRNA synthetase pairs, which in turn mediate the introduction of an ncAA for a certain codon. This approach enables an optimal orthogonality to the autologous translation apparatus and a high flexibility concern-ing the incorporation of multiple ncAAs. As examples for the successful incorporation of non-canonical amino acids, we will develop a rapid test for prions and a new chromatography method for a mild elution of proteins.
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