Searching for the ideal triazole: Investigating the 1,5-triazole as a charge neutral DNA backbone mimic

Abstract

A novel triazole linkage that mimics the phosphodiester backbone in DNA was designed, synthesised and evaluated. Unlike previous work which utilised copper to form a 1,4 triazole linkage in the DNA backbone, a ruthenium catalyst was used to yield a 1,5 triazole. The artificial linkage was incorporated into a DNA backbone via a phosphoramidite building block using solid phase synthesis. The biophysical properties of DNA with a 1,5 triazole linkage in the backbone were evaluated by UV melting and circular dichroism and compared to DNA modified with previously reported 1,4 triazole linkages of various lengths.

A New 1,5-Disubstituted Triazole DNA Backbone Mimic with Enhanced Polymerase Compatibility

“… 32 Destabilizing effects have also been reported for TL 1 , TL 3 , TL 4 , and TL 6 ( Table 1 ). 38 All 6-bond TLs (TLs 2 – 4 ) generally show more favorable duplex stability properties than their shorter 5-bond (TL 1 ) or longer 7-bond (TL 5 and TL 6 ) counterparts. Rigid TL 1 is the most destabilizing among the TLs.…”

“…Despite some destabilizing effects, TL 1 , TL 3 , TL 4 , and TL 6 are known to retain global B- or A-form duplex structures with the DNA or RNA target, respectively. 38 In the current study, duplexes formed by TL 2 - and TL 5 -modified ON2 and ON5 with complementary DNA or RNA were analyzed by circular dichroism (CD) spectroscopy ( Figure 2 ). The duplexes of ON2 with DNA and RNA show almost identical spectra to the equivalent duplexes formed by unmodified ON7.…”

“…Thus, we compared TLs 1 – 7 in primer extension studies ( Figure 3 and Figure 4 ). We synthesized unmodified templates and TL-modified templates, each containing a single TL (TLs 1 – 7 ), using phosphoramidite 6 (TL 2 ), 9 (TL 5 ), or previously reported phosphoramidites (TL 1 , 19 TL 3 , 38 TL 4 , 27 TL 6 , 43 and TL 7 19 ). We then assessed the polymerase compatibilities of TLs 1 – 7 by enzymatic extension of fluorescein (FL)-labeled primers followed by analysis of the FL-labeled products after gel electrophoresis.…”

“… 9 In search of the ideal nucleic acid triazole linkage, we recently developed a 1,5-disubstituted 1,2,3-triazole internucleoside linkage which was prepared by Ru II -catalyzed azide–alkyne cycloaddition (RuAAC) (TL 3 ; Figure 1 ). 38 Altogether, a diverse toolbox of TLs has been reported over the past two decades, all having distinct triazole orientations and linker lengths as indicated in Figure 1 . However, whether the more recent triazole isoforms, such as the 1,5-disubstituted TL 3 , retain compatibility for replication by DNA polymerases had not yet been determined.…”

“…We directly compare the effect of RuAAC vs CuAAC to form TLs from the same precursors (TL 2 vs TL 5 ), and we study the ability of oligonucleotides containing these TLs to form duplexes with a DNA or RNA target in comparison to reported triazoles TL 1 , TL 3 , TL 4 , and TL 6 . 38 Moreover, we test how efficiently polymerases are able to read through TLs 1 – 7 in replication templates. For the first time, we assess replication through a 1,5-disubstituted 1,2,3-triazole (TL 2 and TL 3 ).…”