Nonenzymatic sequence-specific cleavage of duplex DNA via triple-helix formation by homopyrimidine phosphorothioate oligonucleotides.
Phenanthroline was attached covalently to the 5'-terminus of the unmodified and modified (3'-terminal phosphorothioate) oligonucleotide sequences, TTTTTTCTTCTCTTTCC (OP-17 mer) and TTTTTTTCTTCTCTTTCsC (OPRp-17 mer or OPSp-17 mer) via a phosphoramidite bond. Simian virus 40 DNA contains a single target site for these oligonucleotides. In the presence of copper ions, the efficient double-stranded cleavage at 37 degrees C and pH 7.0 was observed by agarose gel electrophoresis. The asymmetric distribution of the cleavage sites on the two strands revealed that the cleavage reaction took place in the minor groove, even though the linker was located in the major groove. Of particular interest are the 3'-terminal phosphorothioate oligonucleotide-phenanthroline derivatives (Rp or Sp), which were found to have cleavage activities of the same order as for the oligonucleotide phenanthroline (OP-17 mer). Furthermore, the OPSp-17 mer was intact after incubation in 10% fetal bovine serum for 24 h, whereas, the OPRp-17 mer was slightly more unstable than the OPSp-17 mer. However, the OP-17 mer was completely degraded. An increased resistance to nucleases has been observed by the introduction of phosphorothioate groups on the 3'-terminus of oligonucleotide-phenanthroline derivatives. This stabilization should help us to design much more efficient chemical recognition enzymes and antisense nucleic acid based anti-viral therapies, which could be used as tools in cellular biology.
Tsukahara
S
S
Department of Industrial Chemistry, Chiba Institute of Technology, Japan.
Suzuki
J
J
Ushijima
K
K
Takai
K
K
Takaku
H
H
D000818:Animals; D001665:Binding Sites; D001769:Blood / Q000378:metabolism; D002417:Cattle; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D009690:Nucleic Acid Conformation; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry / Q000378:metabolism*; D009841:Oligonucleotides / Q000097:blood / Q000737:chemistry* / Q000378:metabolism*; D010618:Phenanthrolines / Q000737:chemistry; D011743:Pyrimidines / Q000737:chemistry; D013539:Simian virus 40 / Q000235:genetics; D013379:Substrate Specificity
D004279:DNA, Viral; D009692:Nucleic Acid Heteroduplexes; D009841:Oligonucleotides; D010618:Phenanthrolines; D011743:Pyrimidines; D004247:DNA