Improving the Efficiency and Orthogonality of Genetic Code Expansion.

The site-specific incorporation of the noncanonical amino acid (ncAA) into proteins via genetic code expansion (GCE) has enabled the development of new and powerful ways to learn, regulate, and evolve biological functions . However, cellular biosynthesis of ncAA-containing proteins with high efficiency and fidelity is a formidable challenge. In this review, we summarize up-to-date progress towards improving the efficiency and orthogonality of GCE and enhancing intracellular compatibility of introduced translation machinery in the living cells by creation and optimization of orthogonal translation components, constructing genomically recoded organism (GRO), utilization of unnatural base pairs (UBP) and quadruplet codons (four-base codons), and spatial separation of orthogonal translation.