Aminoacyl-tRNA synthetase editing preserves the canonical genetic code

Aminoacyl-tRNA synthetases (aaRS) catalyze ATP- dependent covalent coupling of cognate amino acids and tRNAs for ribosomal protein synthesis. The cellular requirements for accurate aminoacylation are high because this reaction defines the genetic code. Yet, some aaRSs are incapable of discriminating against structurally similar amino acids during the synthetic reaction. To keep errors in aminoacyl-tRNA synthesis low these enzymes have evolved additional editing mechanisms. Pre- transfer editing involves hydrolysis of non- cognate aminoacyl-AMP intermediates within the synthetic active site, and can be stimulated by tRNA. Post-transfer editing operates through hydrolysis of misaminoacylated tRNA in a separate dedicated protein domain. The balance between the pre- and post-transfer editing pathways is dictated by kinetic partitioning of aminoacyl-AMP between the aminoacyl transfer step and hydrolysis. The requirement for rapid synthesis of aminoacyl-tRNA within the aaRS synthetic site may have provided an evolutionary driving force for the acquisition of a separate catalytic module committed to proofreading. This emerging view unveils aaRS proofreading as a fortress of canonical translation. Under various stress conditions nonproteinogenic amino acids may accumulate and threaten the accuracy of protein synthesis. We have shown that the prime target for Escherichia coli leucyl-tRNA synthetase (LeuRS) proofreading is norvaline, a nonproteinogenic amino acid that accumulates under microaerobic growth conditions and decreases cell viability when incorporated into the proteome. This contrasts with the generally accepted view in which a key role for LeuRS editing is to prevent the misincorporation of isoleucine at leucine codons. Our detailed kinetic, thermodynamic, structural and in vivo analyses have established that LeuRS discriminates well against isoleucine in the synthetic reaction ; previous contrasting findings appear to have been based on utilization of impure isoleucine samples contaminated with leucine. Accumulation of norvaline may also jeopardize the accuracy of Ile-tRNAIle synthesis. Hence, isoleucine/norvaline substitution in proteins is prevented by the editing activity of isoleucyl-tRNA synthetase. This work has uncovered aaRS translational quality control as a novel part of the adaptive response that protects E. coli cells in rapidly changing oxygen environments.