Sequence-targeted cleavage of nucleic acids by oligo-[.alpha.]thymidylate-phenanthroline conjugates: parallel and antiparallel double helices are formed with DNA and RNA, respectively

Abstract

Oligodeoxynucleotides can be synthesized by using the .alpha. anomers of nucleoside units. Oligo-.alpha.-deoxynucleotides are resistant to nucleases and could be used to regulate gene expression in vivo. Theoretical calculations were carried out to determine the conformational energy of an oligomeric .alpha.-.beta. duplex (dA)5 .cntdot. (dT)5 where the adenosine strand contains natural .beta.-deoxyribonucleotides and the thymidine strand contains synthetic .alpha.-deoxyribonucleotides. These calculations predict that in the more stable B-like conformation the two strands of the double helix should run parallel to each other whereas in the more stable A-like conformation the two strands should adopt an antiparallel orientation. In order to test these predictions 1,10-phenanthroline was covalently attached to the 5''-end of an .alpha.-octathymidylate. In the presence of copper ions and a reducing agent (.beta.-mercaptopropionic acid), the (phenanthroline)2-copper complex generates OH. radicals that cleave phosphodiester bonds in the complementary sequence to which the .alpha.-octathymidylate is bound. By use of a 27mer oligo-.beta.-deoxynucleotide containing an octadeoxyadenylate sequence as a target for the phenanthroline-substituted .alpha.-(dT)8, cleavage was observed on the 5''-side of the (dA)8 sequence, demonstrating that the .alpha.-.beta. DNA-DNA hybrid formed a double helix with parallel orientation of the two strands. The same result was obtained when .alpha.-(dT)8 was bound to .beta.-(dA)n with n = 8 or 10. When a .beta.-oligoriboadenylate was used as a target, cleavage occurred exclusively on the 3''-side of the (rA)8 or (rA)10 sequence, indicating that the .alpha.-.beta. DNA-RNA hybrid formed a double helix with an antiparallel orientation of the two strands. When a phenanthroline-substituted .beta.-octathymidylate was used instead of the .alpha.-octathymidylate, an antiparallel double helix was formed independently of whether the target .beta. sequence was a DNA or an RNA.