The ability of RNA to catalyze RNA ligation is critical to its central role in many prebiotic model scenarios, in particular the copying of information during self‐replication. Prebiotically plausible ribozymes formed from short oligonucleotides can catalyze reversible RNA cleavage and ligation reactions, but harsh conditions or unusual scenarios are often required to promote folding and drive the reaction equilibrium towards ligation. Here, we demonstrate that ribozyme activity is greatly enhanced by charge‐mediated phase separation with poly‐L‐lysine, which shifts the reaction equilibrium from cleavage in solution to ligation in peptide‐RNA coaggregates and coacervates. This compartmentalization enables robust isothermal RNA assembly over a broad range of conditions, which can be leveraged to assemble long and complex RNAs from short fragments under mild conditions in the absence of exogenous activation chemistry, bridging the gap between pools of short oligomers and functional RNAs.

RNA‐catalyzed RNA ligation is widely believed to be a key reaction for primordial biology. However, since typical chemical routes towards activating RNA substrates are incompatible with ribozyme catalysis, it remains unclear how prebiotic systems generated and sustained pools of activated building blocks needed to form increasingly larger and complex RNA. Herein, we demonstrate in situ activation of RNA substrates under reaction conditions amenable to catalysis by the hairpin ribozyme. We found that diamidophosphate (DAP) and imidazole drive the formation of 2′,3′‐cyclic phosphate RNA mono‐ and oligonucleotides from monophosphorylated precursors in frozen water‐ice. This long‐lived activation enables iterative enzymatic assembly of long RNAs. Our results provide a plausible scenario for the generation of higher‐energy substrates required to fuel ribozyme‐catalyzed RNA synthesis in the absence of a highly evolved metabolism.

Catalysis by nucleic acids is indispensable for extant cellular life, and it is widely accepted that nucleic acid enzymes were crucial fort he emergence of primitive life 3.5-4b illion years ago. However,g eochemicalc onditions on early Earth must have differed greatlyf rom the constant internal milieus of today's cells. In order to explore plausible scenarios for early molecular evolution, it is therefore essential to understand how different physicochemical parame-ters, such as temperature,p H, and ionic composition, influence nucleic acidc atalysis and to explore to what extent nucleic acid enzymes can adapt to non-physiological conditions. In this article, we give an overview of the research on catalysis of nucleic acids, in particularc atalytic RNAs (ribozymes) and DNAs( deoxyribozymes), under extreme and/or unusualc onditions that may relate to prebiotic environments.[a] Dr.

The ability of RNA to catalyze RNA ligation is critical to its central role in many prebiotic model scenarios, in particular the copying of information during self‐replication. Prebiotically plausible ribozymes formed from short oligonucleotides can catalyze reversible RNA cleavage and ligation reactions, but harsh conditions or unusual scenarios are often required to promote folding and drive the reaction equilibrium towards ligation. Here, we demonstrate that ribozyme activity is greatly enhanced by charge‐mediated phase separation with poly‐L‐lysine, which shifts the reaction equilibrium from cleavage in solution to ligation in peptide‐RNA coaggregates and coacervates. This compartmentalization enables robust isothermal RNA assembly over a broad range of conditions, which can be leveraged to assemble long and complex RNAs from short fragments under mild conditions in the absence of exogenous activation chemistry, bridging the gap between pools of short oligomers and functional RNAs.