Oligonucleotides with "clickable" sugar residues: Synthesis, duplex stability, and terminal versus central interstrand cross-linking of 2′-O-propargylated 2-aminoadenosine with a bifunctional azide

Abstract

Duplex DNA with terminal and internal sugar cross-links were synthesized by the CuAAC reaction from oligonucleotides containing 2′-O-propargyl-2- aminoadenosine as a clickable site and a bifunctional azide (4). Stepwise click chemistry was employed to introduce cross-links at internal and terminal positions. Copper turnings were used as catalyst, reducing the copper load of the reaction mixture and avoiding complexing agents. For oligonucleotide building block synthesis, a protecting group strategy was developed for 2′-O-propargyl-2-aminoadenosine owing to the rather different reactivities of the two amino groups. Phosphoramidites were synthesized bearing clickable 2′-O-propargyl residues (14 and 18) as well as a 2′- deoxyribofuranosyl residue (10). Hybridization experiments of non-cross-linked oligonucleotides with 2,6-diaminopurine as nucleobase showed no significant thermal stability changes over those containing adenine. Surprisingly, an isobutyryl group protecting the 2-amino function has no negative impact on the stability of DNA-DNA and DNA-RNA duplexes. Oligonucleotide duplexes with cross-linked 2′-O-propargylated 2-aminoadenosine (1) and 2′-O-propargylated adenosine (3) at terminal positions are significantly stabilized (ΔTm = +29 °C). The stability results from a molecularity change from duplex to hairpin melting and is influenced by the ligation position. Terminal ligation led to higher melting duplexes than corresponding hairpins, while duplexes with central ligation sites were less stable. © 2014 American Chemical Society.