Oligonucleotides containing 7-deaza-2-deoxyinosine as universal nucleoside: synthesis of 7-halogenated and 7-alkynylated derivatives, ambiguous base pairing, and dye functionalization by the alkyne - Azide 'click' reaction

Article abstract of DOI:10.1002/hlca.200890131

Oligonucleotides containing 7-deaza-2′-deoxyinosine derivatives bearing 7-halogen substituents or 7-alkynyl groups were prepared. For this, the phosphoramidites 2b - 2g containing 7-substituted 7-deaza-2′-deoxyinosine analogues 1b - 1g were synthesized (Scheme 2). Hybridization experiments with modified oligonucleotides demonstrate that all 2′-deoxyinosine derivatives show ambiguous base pairing, as 2′-deoxyinosine does. The duplex stability decreases in the order C_d>A_d>T _d>G_d when 2b - 2g pair with these canonical nucleosides (Table 6). The self-complementary duplexes 5′-d(F⁷c ⁷I-C)₆, d(Br⁷c⁷I-C)₆, and d(I7c7I-C)6 are more stable than the parent duplex d(c⁷I-C) ₆ (Table 7). An oligonucleotide containing the octa-1,7-diyn-1-yl derivative 1g, i.e., 27, was functionalized with the nonfluorescent 3-azido-7-hydroxycoumarin (28) by the Huisgen - Sharpless - Meldal cycloaddition 'click' reaction to afford the highly fluorescent oligonucleotide conjugate 29 (Scheme 3). Consequently, oligonucleotides incorporating the derivative 1g bearing a terminal C≡C bond show a number of favorable properties: i) it is possible to activate them by labeling with reporter molecules employing the 'click' chemistry. ii) Space demanding residues introduced in the 7-position of the 7-deazapurine base does not interfere with duplex structure and stability (Table 8). iii) The ambiguous pairing character of the nucleobase makes them universal probes for numerous applications in oligonucleotide chemistry, molecular biology, and nanobiotechnology.