Intrastrand backbone-nucleobase interactions stabilize unwound right-handed helical structures of heteroduplexes of L-aTNA/RNA and SNA/RNA

Abstract

Xeno nucleic acids, which are synthetic analogues of natural nucleic acids, have potential for use in nucleic acid drugs and as orthogonal genetic biopolymers and prebiotic precursors. Although few acyclic nucleic acids can stably bind to RNA and DNA, serinol nucleic acid (SNA) and L-threoninol nucleic acid (L-aTNA) stably bind to them. Here we disclose crystal structures of RNA hybridizing with SNA and with L-aTNA. The heteroduplexes show unwound right-handed helical structures. Unlike canonical A-type duplexes, the base pairs in the heteroduplexes align perpendicularly to the helical axes, and consequently helical pitches are large. The unwound helical structures originate from interactions between nucleobases and neighbouring backbones of L-aTNA and SNA through CH-O bonds. In addition, SNA and L-aTNA form a triplex structure via C:G*G parallel Hoogsteen interactions with RNA. The unique structural features of the RNA-recognizing mode of L-aTNA and SNA should prove useful in nanotechnology, biotechnology, and basic research into prebiotic chemistry.

Fig. 1. Dimer of duplex structures of L-aTNA/RNA and SNA/RNA stabilized by Hoogsteen base pairs.

Base-pairing patterns and structures of (a) L-aTNA/RNA and (b) SNA/RNA. In the sequence schematics, Watson–Crick and Hoogsteen base pairs are indicated as black circles and dashed lines, respectively. In the stick representations, carbon atoms are coloured magenta, cyan, and green, nitrogens are in blue, oxygens are in red, and phosphorus is in orange. 2F_o-F_c electron density maps contoured at 1.0 σ level. The structures were deposited in the Protein Data Bank. PDB accession codes: 7BPF (L-aTNA/RNA) and 7BPG (SNA/RNA).

Fig. 2. Canonical Watson–Crick and parallel-type Hoogsteen base pairing patterns of L-aTNA/RNA and SNA/RNA.

Watson–Crick (a) and Hoogsteen (b) interactions in _LT8a/R8Br and S8a/R8Br. C, N, O, and P atoms are coloured as in Fig. 1. 2F_o-F_c electron density maps contoured at 1.0 σ level. Arrows indicate relative polarities of the backbones of RNA, L-aTNA, and SNA.

Fig. 3. Dimers of L-aTNA/RNA and of SNA/RNA duplexes connected by end-to-end stacking pack together in crystals.

a Stick representations showing end-to-end stacking of two L-aTNA/RNA duplexes. Top view of the end-to-end stacking between terminal base pairs is also indicated. b High-order helical structure of two dimers of the L-aTNA/RNA duplex. c Packing observed in the crystal of L-aTNA/RNA. d Stick representations showing end-to-end stacking of SR-1 and SR-2 SNA/RNA duplex. Top view of the end-to-end stacking between terminal base pairs is also indicated. e Packing observed in the crystal of SNA/RNA. Carbon atoms are coloured magenta, pink, cyan, sky blue, green, dark green, yellow, and olive. N, O, and P atoms are coloured as in Fig. 1.

Fig. 4. Structures of L-aTNA/RNA and SNA/RNA are unwound in comparison to A-type duplex structure.

a Superposition of duplex structures of L-aTNA/RNA (cyan) and SNA/RNA (green), dsRNA (magenta), LNA/RNA (orange), or PNA/RNA (purple) are shown. PDB accession codes were the following: RNA/RNA, 3ND4; LNA/RNA, 1H0Q; PNA/RNA, 5EMF. b Surface views of minor and major groove of L-aTNA/RNA and SNA/RNA. An example of location of CH₃ on L-aTNA backbone is shown as dotted circle. Carbon atoms are coloured white and N, O, and P atoms are coloured as in Fig. 1.

Fig. 5. Comparison of backbone torsion angles of RNA, L-aTNA, and SNA.

a–c Chemical structures of nucleotides and stick representations in the context of a duplex with RNA for RNA (a), L-aTNA (b), and SNA (c). Carbon atoms are shown in white, cyan, and green for RNA, L-aTNA, and SNA, respectively. N, O, and P atoms are coloured as in Fig. 1.

Fig. 6. Intrastrand hydrogen bonds between backbone carbonyl oxygens and neighbouring nucleobases.

a Short C–O distance within the van der Waals distance cutoff of 3.7 Å observed in L-aTNA/RNA and SNA/RNA are indicated as dashed line in black. Hydrogen bonds between backbone and water are also indicated as dashed line in magenta. b Close up view of C–O interactions of O5′ (backbone)–C8 (adenine), O5'–C6' (backbone), and O2 (cytosine)–C6′ and hydrogen bonds of amide (backbone)–water molecule and O3′–water molecule. c Close up view of C–O interactions of O5′–C6 (cytosine) and O5′–C5 (cytosine) at terminal cytosine residue in SNA. Carbon atoms are shown in white, cyan, and green for RNA, L-aTNA and SNA, respectively. N, O, and P atoms are coloured as in Fig. 1.

Fig. 7. Dimer formation of L-aTNA/RNA, SNA/RNA, L-aTNA/ L-aTNA, and SNA/SNA duplexes in solution.

Mass spectrum of solution of _LT8a and R8Br (a), S8a and R8Br (b), _LT8a and _LT7b (c), and S8a and S7b (d) were collected under non-denaturing conditions in negative ionization mode. Calculated masses are summarized in materials and methods.