Synthesis and characterization of oligodeoxynucleotides containing O6-methyl-, O6-ethyl-, and O6-isopropylguanine

Abstract

The carcinogenicity and mutagenicity of N-nitroso compounds is believed to result primarily from alkylation of DNA on O6 of guanine. To study the base-pairing properties of O6-alkylguanines and the structural changes produced in DNA by their presence, self-complementary dodecanucleotides were synthesized of the general structure CGCXAGCTYGCG (where X was O6-methylguanine, O6-ethylguanine, or O6-isopropylguanine and Y one of the natural bases). The O6-alkyldeoxyguanosines used as building blocks for the synthesis of the oligomers were prepared from the 6-(2-mesitylenesulfonyl) derivative by successive displacement with N-methylpyrrolidine and then alkoxide ions. The difficulties previously encountered in the synthesis of oligomers containing O6-alkylguanine [Borowsy-Borowski, H., & Chambers, R. W. (1987) Biochemistry 26, 2465-2471] were overcome by use of phenylacetyl as N2-protection for the O6-alkylguanine residue. The lability of the phenylacetyl group allowed very mild treatment with ammonia to be used for deprotection, and there was no detectable contamination of the product with 2,6-diaminopurine. Depurination of O6-alkylguanine residues with a free 5′-OH, which can occur if a protic acid is used to remove the 5′-protecting group from the alkylated nucleoside during oligonucleotide synthesis, was avoided by using ZnBr2. The oligonucleotide sequences that would form double helices containing O6-alkylG·C base pairs had lower optical melting temperatures (Tm) than the parent with G·C pairs, but only the isopropylG·C oligomer had significantly less hypochromicity. But the sequences giving helices with O6-alkylG·T base pairs had less hypochromicity than the parent and had biphasic melting profiles that began at a significantly lower temperature than the sequences with O6-alkylG·C base pairs. © 1989 American Chemical Society.