Year: 2021
Type: article
Abstract: Genetic code expansion technologies supplement the natural codon repertoire with assignable variants in vivo, but are often limited by heterologous translational components and low suppression efficiencies. Here, we explore engineered Escherichia coli tRNAs supporting quadruplet codon translation by first developing a library-cross-library selection to nominate quadruplet codon-anticodon pairs. We extend our findings using a phage-assisted continuous evolution strategy for quadruplet-decoding tRNA evolution (qtRNA-PACE) that improved quadruplet codon translation efficiencies up to 80-fold. Evolved qtRNAs appear to maintain codon-anticodon base pairing, are typically aminoacylated by their cognate tRNA synthetases, and enable processive translation of adjacent quadruplet codons. Using these components, we showcase the multiplexed decoding of up to four unique quadruplet codons by their corresponding qtRNAs in a single reporter. Cumulatively, our findings highlight how E. coli tRNAs can be engineered, evolved, and combined to decode quadruplet codons, portending future developments towards an exclusively quadruplet codon translation system.
Source: Nature Communications
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $5,560
Funders: U.S. Department of Health and Human Services, National Aeronautics and Space Administration, Broad Institute, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, National Institutes of Health, NIH Office of the Director, National Institute of General Medical Sciences, National Institute of Allergy and Infectious Diseases
OpenAlex ID: W3202524566