Template-Directed Nonenzymatic Primer Extension Using 2-Methylimidazole-Activated Morpholino Derivatives of Guanosine and Cytidine.

Abstract

Efforts to develop self-replicating nucleic acids have led to insights into the origin of life and have also suggested potential pathways to the design of artificial life forms based on non-natural nucleic acids. The template-directed nonenzymatic polymerization of activated ribonucleotide monomers is generally slow because of the relatively weak nucleophilicity of the primer 3′-hydroxyl. To circumvent this problem, several nucleic acids based on amino-sugar nucleotides have been studied, and as expected, the more-nucleophilic amine generally results in faster primer extension. Extending this logic, we have chosen to study morpholino nucleic acid (MoNA), because the secondary amine of the morpholino-nucleotides is expected to be highly nucleophilic. We describe the synthesis of 2-methylimidazole-activated MoNA monomers from their corresponding ribonucleoside 5′-monophosphates and the synthesis of an RNA primer with a terminal MoNA nucleotide. We show that the activated G and C MoNA monomers enable rapid and efficient extension of the morpholino-terminated primer on homopolymeric and mixed-sequenced RNA templates. Our results show that MoNA is a non-natural informational polymer that is worthy of further study as a candidate self-replicating material.