Double-Headed 2'-Deoxynucleotides That Hybridize to DNA and RNA Targets via Normal and Reverse Watson-Crick Base Pairs.

Through the use of modified nucleotides, synthetic nucleic acids have found several fields of application within biotechnology and in the pharmaceutical industry. We have previously introduced nucleotides with an additional functional nucleobase linked to C2' of arabinonucleotides (BX). These double-headed nucleotides fit neatly into DNA·DNA duplexes, where they can replace the corresponding natural dinucleotides and thus condense the molecular information. Here, we introduce a 2'-deoxy version of the BX design with inversion of the C2' stereochemistry (dSBX) with the aim of obtaining improved RNA recognition. Specifically, dSBX analogues with cytosine or isocytosine attached to C2' of 2'-deoxyuridine (dSUC and dSUiC) were synthesized and evaluated in duplexes. Whereas the dSBX design did not outperform the BX design in terms of mimicking dinucleotides in nucleic acid duplexes, it was able to engage in reverse Watson-Crick pairing using its 2'-base. This was evident from the ability of the dSUC cytosine to form stable mis-matching base pairs with opposite cytosines identified as hemiprotonated C·C+ pairs. Furthermore, specific base-pairing with guanine was only observed for the isocytosine-bearing dSUiC monomer. Very stable duplexes were obtained with dSUC/iC monomers in each strand indicating that fully modified double-headed nucleic acid sequences could be based on the dSBX design.

Double-headed nucleotides as xeno nucleic acids: information storage and polymerase recognition.

Xeno nucleic acids (XNAs) are artificial genetic systems based on sugar-modified nucleotides. Herein, we investigate double-headed nucleotides as a new XNA. A new monomer, AT, is presented, and together with previous double-headed nucleotide monomers, new nucleic acid motifs consisting of up to five consecutive A·T base pairs have been obtained. Sections composed entirely of double-headed nucleotides are well-tolerated within a DNA duplex and can condense the genetic information. For instance, a 13-mer duplex is condensed to an 11-mer modified duplex containing four double-headed nucleotides while simultaneously improving duplex thermal stability with +14.0 °C. Also, the transfer of information from double-headed to natural nucleotides by DNA polymerases has been examined. The first double-headed nucleoside triphosphate was prepared but could not be recognized and incorporated by the tested DNA polymerases. On the other hand, it proved possible for Therminator DNA polymerase to transfer the information of a double-headed nucleotide in a template sequence to natural DNA under controlled conditions.

Double-headed nucleic acids condense the molecular information of DNA to half the number of nucleotides.

Nucleotide monomers that hold two nucleobases each, i.e. double-headed nucleotides, have been shown to form two sets of functional Watson-Crick base pairs when incorporated into dsDNA, and they hereby behave as dinucleotides. To form the basis for fully modified double-headed nucleic acids (DhNA), we have prepared three new DhNA monomers and can now demonstrate that the molecular information of 10 Watson-Crick base pairs can be condensed to highly stable 5-mer DhNA duplexes.