Prebiotically Plausible RNA Activation Compatible with Ribozyme-Catalyzed Ligation.

Emilie Yeonwha Song,Huacan Lin,Eddy Ivanhoe Jiménez,Kristian Le Vay,Ramanarayanan Krishnamurthy,Hannes Mutschler

doi:10.1002/anie.202010918

Abstract

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.

Highlights

Emilie Yeonwha Song, Eddy Ivanhoe JimØnez+, Huacan Lin+, Kristian Le Vay, Ramanarayanan Krishnamurthy,* and Hannes Mutschler*
To monitor the coupled activation–ligation reactions, we developed a reporter electrophoretic mobility shift assay (EMSA) based on a cis-ligating version of the hairpin ribozyme (HPz)
We identified the inclusion of 5 mm DAP, 5 mm Mg2+, and 5 mm imidazole as optimal conditions far for the combined activation and ligation of the RNA substrate sub-P (Figure S19), and used these conditions in all further experiments

Summary

Introduction

Emilie Yeonwha Song, Eddy Ivanhoe JimØnez+, Huacan Lin+, Kristian Le Vay, Ramanarayanan Krishnamurthy,* and Hannes Mutschler*. The higher initial concentration of the two components reduces the amount of water-ice necessary to reach the molal equilibrium concentrations of the unfrozen phase.[24] the resulting cHPz and substrate concentrations in the aqueous phase may be lower than with decreased DAP and imidazole concentrations, leading to less efficient activation and/or ligation.

Results

Conclusion