Synthesis of 2′-Deoxy-α‑l‑threofuranosyl Nucleoside Triphosphates

α-l-Threofuranosyl nucleic acid (TNA) is an artificial
genetic polymer in which the natural five-carbon ribose sugar found
in RNA has been replaced with an unnatural four-carbon threose sugar.
Despite a different sugar–phosphate backbone, TNA is capable
of forming stable, antiparallel Watson–Crick duplex structures
with itself and with complementary strands of DNA and RNA. This property
of intersystem base pairing, coupled with the chemical simplicity
of threose relative to ribose, provides support for TNA as a candidate
RNA progenitor in the evolution of life. In an effort to evaluate
the functional properties of TNA by in vitro evolution, engineered
polymerases have been developed that are capable of copying information
back and forth between DNA and TNA. However, the current generation
of TNA polymerases function with reduced activity relative to their
natural counterparts, which limits the evaluation of TNA as a primordial
genetic material. Here, we describe the chemical synthesis and polymerase
recognition of 2′-deoxy-α-l-threofuranosyl nucleoside
3′-triphosphates (dtNTPs) as chain-terminating reagents in
a polymerase-mediated TNA synthesis reaction. The synthesis of dtNTPs
should make it possible to investigate the mechanism of TNA synthesis
by X-ray crystallography by trapping the polymerase in the catalytically
active conformation.