Recent advances in genetic code engineering in Escherichia coli

The expansion of the genetic code is gradually becoming a core discipline in Synthetic Biology. It offers the best possible platform for the transfer of numerous chemical reactions and processes from the chemical synthetic laboratory into the biochemistry of living cells. The incorporation of biologically occurring or chemically synthesized non-canonical amino acids into recombinant proteins and even proteomes via reprogrammed protein translation is in the heart of these efforts. Orthogonal pairs consisting of aminoacyl-tRNA synthetase and its cognate tRNA proved to be a general tool for the assignment of certain codons of the genetic code with a maximum degree of chemical liberty.Here, we highlight recent developments that should provide a solid basis for the development of generalist tools enabling a controlled variation of chemical composition in proteins and even proteomes. This will take place in the frame of a greatly expanded genetic code with emancipated codons liberated from the current function or with totally new coding units.

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► Highlights incorporation of industrially relevant non-canonical amino acids.
► Multi-labeling of proteins with distinct non-canonical amino acids.
► Recent developments of new orthogonal pairs.
► Release factor 1 knockout strains for amber codon reassignment.
► Codon emancipation for proteome-wide integration of new chemistries into living cells.