Figure 1.

Schematic illustration of the general principles for the site-specific incorporation of ncAA into proteins in vivo. The natural translational system involves the activity of endogenous synthetase, which charges the endogenous tRNA with natural amino acids (NAAs), which is presented as a red oval shape. The orthogonal translational system is applied to site-specifically incorporate ncAA into proteins. Orthogonal aminoacyl-tRNA synthetase acylates the orthogonal tRNA with a noncanonical amino acid. The acylated orthogonal tRNA introduces the ncAA at a specific site complementary to the unique codon, thereby incorporating it into the ORF that encodes the protein of interest (ncAA is represented by the red star).