DNA with Branched Internal Side Chains: Synthesis of 5-Tripropargylamine-dU and Conjugation by an Azide-Alkyne Double Click Reaction

Abstract

5-Tripropargylamine-2'-deoxyuridine (1a) containing two terminal triple bonds was synthesized by a Pd-assisted Sonogashira cross-coupling reaction and was subsequently converted into the corresponding phosphoramidite building block (9) and employed in solid-phase oligonucleotide synthesis. T-m experiments demonstrated that the presence of covalently attached branched tripropargylamine residues has a positive effect on the base pair stability. The two terminal C C bonds of modified DNA were function alized by means of Cu-1-mediated 1,3-dipolar cycloaddition reactions (click chemistry) with azides such as 3-azido-7-hydroxycoumarin or 3-azido-3'- deoxythymidine (AZT) both in solution and on solid support in particular, with the nonfluorescent nonfluorescent 3-azido-7-hydroxycoumarin a strongly fluorescent oligonucleotide bis-dye conjugate was generated. For comparison, the N(3)-propargylated 2-deoxyuridine 2 was prepared from 2'-deoxyuridine and propargyl bromide and incorporated into DNA. The two terminal pargylamine triple bonds of 1a allow the simultaneous post-modification of DNA by two reporter molecules and can be applied to almost any azido derivatives (oligonucleotides, proteins, polysaccharides etc.) including those forming dendrimeric side chains.