Translation‐Promoting Effects of RNA Template Overhangs in the Absence of Ribosomes

Abstract Translation requires a complex set of biomacromolecules. How it evolved is an unsolved problem. Ribosome‐free, single‐nucleotide translation via coupling of 3′‐aminoacylated RNA to 5′‐phosphoramidate‐linked RNAs was shown for very short peptides. It was unclear how this process could be induced at a specific locus, regulated, and expanded. Here we show that a triplex‐forming template overhang accelerates ribosome‐free translation and increases its yield. An NMR‐monitored model study suggested that mixed anhydrides between carboxylates of amino acid residues and phosphates can contribute to efficient peptide coupling, strengthening proximity effects. Folding into triplexes at pH 6 enhances the template effect, and unfolding at pH 8 suppresses it, suggesting a simple way of achieving “inducible translation” in a system devoid of biomacromolecules. Under mild acidic conditions, doubly RNA‐linked peptides up to octamers were formed in near‐quantitative yield. Taken together, our data suggests that the growth of template strands not only allows for encoding more genetic information, but may also enhance translational fitness.