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 (1 a) 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 demonstrate that the presence of covalently attached branched tripropargylamine residues has a positive effect on the base pair stability. The two terminal C[triple chemical bond]C bonds of modified DNA were functionalized by means of Cu(I)-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 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 triple bonds of 1 a 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.