Engineering aminoacyl-tRNA synthetases for use in synthetic biology.

ABSTRACT

Within the broad field of synthetic biology, genetic code expansion (GCE) techniques enable creation of proteins with an expanded set of amino acids. This may be invaluable for applications in therapeutics, bioremediation, and biocatalysis. Central to GCE are aminoacyl-tRNA synthetases (aaRSs) as they link a non-canonical amino acid (ncAA) to their cognate tRNA, allowing ncAA incorporation into proteins on the ribosome. The ncAA-acylating aaRSs and their tRNAs should not cross-react with 20 natural aaRSs and tRNAs in the host, i.e., they need to function as an orthogonal translating system. All current orthogonal aaRS•tRNA pairs have been engineered from naturally occurring molecules to change the aaRS's amino acid specificity or assign the tRNA to a liberated codon of choice. Here we discuss the importance of orthogonality in GCE, laboratory techniques employed to create designer aaRSs and tRNAs, and provide an overview of orthogonal aaRS•tRNA pairs for GCE purposes.