Uniform affinity-tuning of N-methyl-aminoacyl-tRNAs to EF-Tu enhances their multiple incorporation

Yoshihiko Iwane, Hiroyuki Kimura, Takayuki Katoh, and Hiroaki Suga

Nucleic Acids Research, gkab288, https://doi.org/10.1093/nar/gkab288

In the ribosomal peptide synthesis, all kinds of proteinogenic aminoacyl-tRNAs (AA-tRNAs) are delivered by EF-Tu to the ribosome with similar efficiencies, which is achieved by the uniform binding affinities between those AA-tRNAs and EF-Tu. In contrast, N-methyl amino acids (MeAAs) are poorly incorporated into peptides because EF-Tu weakly binds to MeAA-tRNAs compared to the proteinogenic AA-tRNAs. To overcome this issue, the T-stem region of MeAA-tRNAs was substituted with a designer sequence, resulting in the reinforcement of their EF-Tu affinities to the comparable level with those of proteinogenic AA-tRNAs. This uniform affinity-tuning methodology has enabled the incorporation of nine distinct MeAAs into linear and thioether-macrocyclic model peptides.

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