Replacing uridine with 2-thiouridine enhances the rate and fidelity of nonenzymatic RNA primer extension.

Abstract

The nonenzymatic replication of RNA oligonucleotides is thought to have played a key role in the origin of life prior to the evolution of ribozyme-catalyzed RNA replication. Although the copying of oligo-C templates by 2-methylimidazole-activated G monomers can be quite efficient, the copying of mixed sequence templates, especially those containing A and U, is particularly slow and error-prone. The greater thermodynamic stability of the 2-thio-U(s2U):A base pair, relative to the canonical U:A base pair, suggests that replacing U with s2U might enhance the rate and fidelity of the nonenzymatic copying of RNA templates. Here we report that this single atom substitution in the activated monomer improves both the kinetics and the fidelity of nonenzymatic primer extension on mixed-sequence RNA templates. In addition, the mean lengths of primer extension products obtained with s2U is greater than those obtained with U, augmenting the potential for nonenzymatic replication of heritable function-rich sequences. We suggest that noncanonical nucleotides such as s2U may have played a role during the infancy of the RNA world by facilitating the nonenzymatic replication of genomic RNA oligonucleotides.