Linkage structures strongly influence the binding cooperativity of DNA intercalators conjugated to triplex forming oligonucleotides

摘要

Conjugation of DNA intercalators to triple heiix forming oiigodeoxynucieotides (ODN's) can enhance ODN binding properties and consequentiy their potentiai abiilty to moduiate gene expression. To test the hypothesis that iinkage structure couid strongiy Infiuence the binding enhancement of intercaiator conjugation with triplex forming ODN's, we have used a modei system to investigate binding avidity of short oiigomers conjugated to DNA intercaiators through various linkages. Using a dA₁₀·T₁₀ target sequence imbedded in a 20 bp duplex, binding avidities of a T₁₀ ODN joined to the DNA intercalator 6,9-diamino, 3-methoxy acridine (DAMA) by 8 different 5′ linkages were measured using an electrophoretic mobiiity shift assay. Afthough unmodified T₁₀ has a very limited capacity for stabie binding under these conditions (apparent K_d 250 μM at 4°C), conjugation to DAMA using tiexible linkers of certain lengths and chemicai compositions greatly enhanced binding (K_d of 1 μM at 4°C). Other linkers, however, modestiy enhanced binding or had no effect on binding at au. Thus, the length, flexibility, and chemicai composition of iinker structures au substantialiy influence intercalator conjugated oilgodeoxynucleotide binding avidity.