Site-specific incorporation of non-natural residues into peptides: Effect of residue structure on suppression and translation efficiencies

Slow peptide bond formation by proline and other N -alkylamino acids in translation

“…N-alkyl groups in polypeptides and proteins kink 3-dimensional structures, result in slow isomerization between cis and trans conformations in the case of Pro, and can improve the pharmacological properties of peptides by decreasing their degradation by proteases and increasing their membrane permeability. To investigate these special properties or endow them on translation products, there have been many studies incorporating N-alkylamino acids (''imino acids'') in translation (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13). Small linear N-alkylamino acids ( Fig.…”

“…Small linear N-alkylamino acids ( Fig. 1 Upper Right), because of their similarity to Pro, were expected to incorporate well in translation, but the efficiencies were variable (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13). Incomplete incorporations in purified translation systems (8)(9)(10)(11)(12)(13) are particularly puzzling because of the lack of competing reactions, incubation times of tens of minutes, and an average codon translation time comparable with that in Escherichia coli (45 ms) (14).…”

Posttranslational Modification of Proteins Incorporating Nonnatural Amino Acids

“…The suppression efficiency was 65% for 125 I-Tyr; in the case of the other NAAs, it was 72% for N-methylphenylalanine and 46% for phenyllactic acid. Using this approach, a variety of NAAs have been incorporated into proteins in vitro with suppression efficiencies of up to 72% [42]. Chemical Aminoacylation of tRNA, Four-Base Codon Of the 64 codons, all except for the three stop codons are assigned to a particular natural amino acid.…”

Optimal Codon Choice Can Improve the Efficiency and Fidelity of N‐Methyl Amino Acid Incorporation into Peptides by In‐Vitro Translation

“…[1–11] However, N -Me AAs are suboptimal substrates for the translational apparatus, [12] and as a result, premature termination of translation can occur or incorrect amino acids can be misincorporated in place of the desired N -Me AA, resulting in reduced incorporation efficiency and fidelity of the N -Me AA. We sought to determine the main cause of such misincorporation and truncation events, and then to optimize the incorporation of N -Me AAs.…”