Modification of DNA with octadiynyl side chains: Synthesis, base pairing, and formation of fluorescent coumarin dye conjugates of four nucleobases by the alkyne-azide ``click'' reaction

Abstract

Oligonucleotides incorporating 5-(octa-1,7-diynyl)-2'-deoxycytidine 1a, 5-(octa-1,7-diynyl)-2'-deoxyuridine 2a and 7-deaza-7-(octa-1,7-diynyl)-2'-deoxyguanosine 3a, 7-deaza-7-(octa-1,7-diynyl)-2'-deoxyadenosine 4a were prepared. For this, the phosphoramidites 7, 10, and 13 were synthesized and employed in solid-phase oligonucleotide synthesis. The octa-1,7-diynyl nucleosides 1a-4a were obtained from their corresponding iodo derivatives using the palladium-assisted Sonogashira cross-coupling reaction. The Tm values demonstrated that DNA duplexes containing octa-1,7-diynyl nucleosides show a positive influence on the DNA duplex stability when they are introduced at the 5-position of pyrimidines or at the 7-position of 7-deazapurines. The terminal alkyne residue of oligonucleotides were selectively conjugated to the azide residue of the nonfluorescent 3-azido-7-hydroxycoumarin (38) using the protocol of copper(I)-catalyzed [3 2] Huisgen-Sharpless-Meldal cycloaddition ``click chemistry'' resulting in the formation of strongly fluorescent 1,2,3-triazole conjugates. The fluorescence properties of oligonucleotides with covalently linked coumarin-nucleobase assemblies were investigated. Among the four modified bases, the 7-deazapurines show stronger fluorescence quenching than that of pyrimidines.