Synthesis and characterization of oligodeoxynucleotides containing 4-O-methylthymine.

The carcinogenicity of N-nitroso compounds is believed to result from the alkylation of DNA, particularly on O-6 of the guanine and O-4 of the thymine residues. In order to study the base-pairing properties of 4-O-methylthymidine (T*) residues and the structural changes produced in DNA by the presence of this alkylated nucleoside, the oligodeoxyribonucleotides TGCG, CGCAAGCTTGCG, CGCGAGCTTGCG, and CGCAAGCTTGCG were synthesized by the phosphotriester approach in solution. The 4-O-methylthymidine required for oligonucleotide synthesis was prepared by treating the 4-(3-nitro-1,2,4-triazolo) derivative of 3',5'-bis-O-(methoxyacetyl)thymidine with 1,8-diazabicyclo[5.4.0]-undec-7-ene (DBU) in methanol solution. The susceptibility of the 4-O-methyl group of T toward nucleophiles enables this group of 4-O-methylthymidine-containing oligomers to be labeled by a direct exchange reaction with [13C]- or [14C]methanol in the presence of DBU. Although it has been previously suggested that 4-O-methylthymine forms stable base pairs with guanine, the thermal melting profiles of the double helices formed by these dodecamers suggest that the presence of 4-O-methylthymine paired to either adenine or guanine destablizes the helix. The melting curve of the sequence containing a 4-O-methylthymine residue base paired to guanine was biphasic and similar to that of an analogous sequence containing 6-O-methylguanine paired to thymine.