Fast and accurate nonenzymatic copying of an RNA-like synthetic genetic polymer

Abstract

Recent advances suggest that it may be possible to construct simple artificial cells from two subsystems: a self-replicating cell membrane and a self-replicating genetic polymer. Although multiple pathways for the growth and division of model protocell membranes have been characterized, no self-replicating genetic material is yet available. Nonenzymatic template-directed synthesis of RNA with activated ribonucleotide monomers has led to the copying of short RNA templates; however, these reactions are generally slow (taking days to weeks) and highly error prone. N3α- P5α-linked phosphoramidate DNA (3α-NP-DNA) is similar to RNA in its overall duplex structure, and is attractive as an alternative to RNA because the high reactivity of its corresponding monomers allows rapid and efficient copying of all four nucleobases on homopolymeric RNA and DNA templates. Here we show that both homopolymeric and mixed-sequence 3α-NP-DNA templates can be copied into complementary 3α-NP-DNA sequences. G:T and A: C wobble pairing leads to a high error rate, but the modified nucleoside 2-thiothymidine suppresses wobble pairing. We show that the 2-thiothymidine modification increases both polymerization rate and fidelity in the copying of a 3α-NP-DNA template into a complementary strand of 3α-NP-DNA. Our results suggest that 3α-NP-DNA has the potential to serve as the genetic material of artificial biological systems.