RNA Oligomerisation without Added Catalyst from 2 ',3 '-Cyclic Nucleotides by Drying at Air-Water Interfaces

For the emergence of life, the abiotic synthesis of RNA from its monomers is a central step. We found that in alkaline, drying conditions in bulk and at heated air-water interfaces, 2 ',3 '-cyclic nucleotides oligomerised without additional catalyst, forming up to 10-mers within a day. The oligomerisation proceeded at a pH range of 7-12, at temperatures between 40-80 degrees C and was marginally enhanced by K+ ions. Among the canonical ribonucleotides, cGMP oligomerised most efficiently. Quantification was performed using HPLC coupled to ESI-TOF by fitting the isotope distribution to the mass spectra. Our study suggests a oligomerisation mechanism where cGMP aids the incorporation of the relatively unreactive nucleotides C, A and U. The 2 ',3 '-cyclic ribonucleotides are byproducts of prebiotic phosphorylation, nucleotide syntheses and RNA hydrolysis, indicating direct recycling pathways. The simple reaction condition offers a plausible entry point for RNA to the evolution of life on early Earth

High-Fidelity Templated Ligation of RNA via 2′,3′-cyclic Phosphate

The templated ligation of oligonucleotides offers a mode of replication in an RNA world. The 2′,3′-cyclic phosphate (>P) is a prebiotically available activation group for RNA and the product of backbone hydrolysis. Using gel electrophoresis and liquid chromatography, we found that the templated ligation of RNA with >P activation proceeds in alkaline (pH 9-11) low-salt aqueous solutions with 1 mM MgCl2 in temperatures ranging from 20 to 25 °C within a few days. Under the optimum conditions of pH 10 and 5 °C, the ligation yielded 40% after 7 days. No additional catalysts were required. In contrast to previous reports, we found an equimolar mixture of 2′-5′ and 3′-5′ linked oligomers in the used conditions. We probed the nucleotide specificity at the ligation site and found that one mutation reduced the ligation yield by 82-92%. We extrapolated these results to a per-nucleotide replication fidelity of 95-98% when ligating 4- to 6-mers. With splinted oligomers, five ligations created a 96 mer strand, demonstrating a possible assembly pathway for long ribozymes. With the low salt requirements, strand separation will be compatible with the ligation conditions using non-equilibrium settings. The findings suggest that templated ligation mediated by 2′,3′-cyclic phosphate in alkaline conditions offer a slow, but precise replication and elongation reaction for RNA on early Earth

RNA polymerisation without catalyst from 2’,3’-cyclic nucleotides by drying at air-water interfaces

For the emergence of life, the abiotic synthesis of RNA from its monomers is a central step. We found alkaline, uncatalysed drying conditions in bulk and at heated air-water interfaces where 2´,3´-cyclic nucleotides polymerised, forming up to 10-mers within a day. The polymerisation proceeded at a pH range of 7-12 at temperatures between 40-80 °C and was marginally enhanced by K+ ions. Among the canonical ribonucleotides, cGMP polymerised most efficiently. Quantification was performed using HPLC coupled to ESI-TOF by fitting the isotope distribution to the mass spectra. Our study suggests a polymerisation mechanism where cGMP aids the incorporation of the relatively unreactive nucleotides C, A and U. The 2´,3´-cyclic nucleotides are byproducts of prebiotic phosphorylation, nucleotide syntheses and RNA hydrolysis, indicating direct recycling pathways. The simple reaction condition offers a plausible entry point for RNA to the evolution of life on early Earth