An approach to therapeutic agents through selective targeting of destabilised nucleic acid duplex sequences

The binding of Delta Delta/Delta Delta-[{Ru(phen)(2)}(2)(mu-bb(n))](4+) {where phen = 1,10-phenanthroline, bb(n) = 1,n-bis[4(4'-methyl-2,2'-bipyridyl)]-alkane (Delta Delta/Delta Delta-Rubb(n))} to the non-self complementary oligonucleotide 5'-d (CGCGATAAGCCGC.5'-GCGGCATTACGCG) (3-DB) has been examined using a 4', 6-diamidino-2-phenylindole dihydrochloride (DAPI) displacement assay. The 3-DB oligonucleotide contains two single adenine bulge nucleotides that are separated by three base pairs. H-1 NMR spectroscopy data demonstrated that the adenine bases are intra-helical and that the segment containing the two bulge nucleotides and the three A.T base pairs between the bulges forms a destabilised segment within the stable duplex oligonucleotide. The DAPI displacement assay demonstrated that Delta Delta-Rubb(7)-bound 3-DB with higher affinity than the other members of the Delta Delta/Delta Delta-Rubb(n) series. Molecular models suggested that the seven-carbon chain length in Delta Delta-Rubb(7) was ideal to span the distance between the two bulge sites. The binding of Delta Delta-Rubb(7) to 3-DB was also studied by 1H NMR spectroscopy and molecular modelling. The selective changes in chemical shifts for the resonances from 3-DB upon addition of Delta Delta-Rubb(7) suggested that the metal complex specifically bound at the destabilised segment between A(5) and A(19). Observation in NOESY spectra of NOE cross peaks between 3-DB and Delta Delta-Rubb(7) confirmed that one of the ruthenium centres bound at the A(5) bulge site, with the other metal centre positioned at the A(19) bulge. In addition, Delta Delta-Rubb(7) was found to bind chromosomal DNA extracted from a suspension of Staphylococcus aureus that had been incubated with the ruthenium(II) complex. As inert dinuclear ruthenium(II) complexes are capable of being transported into a bacterial cell and bind chromosomal DNA, it is possible that they could be developed into anti-microbial agents that specifically target destabilised segments of DNA that are recognised by essential DNA-binding proteins.