Solution structure of a wedge-shaped synthetic molecule at a two-base bulge site in DNA

The solution structure of the complex formed between an oligonucleotide containing a two-base bulge (5'-CACGCAGTTCGGAC(.)5'(.)GTCCGATGCGTG) and DDI, a designed synthetic agent, has been elucidated using high-resolution NMR spectroscopy and restrained molecular dynamic simulation. DDI, which has been found to modulate DNA strand slippage synthesis by DNA polymerase I [Kappen, L. S., Xi, Z., Jones, G. B., and Goldberg, I. H. (2003) Biochemistry 42, 2166-2173], is a wedge-shaped spirocyclic molecule whose aglycone structure closely resembles that of the natural product, NCSi-gb, which strongly binds to an oligonucleotide containing a two-base bulge. Changes in chemical shifts of the DNA upon complex formation and intermolecular NOES between DDI and the bulged DNA duplex indicate that agent specifically binds to the bulge site of DNA. The benzindanone moiety of DDI intercalates via the minor groove into the G7-T8-T9(.)A20 pocket, which consists of a helical base pair and two impaired bulge bases, stacking with the G7 and A20 bases. On the other hand, the dihydronaphthalenone and aminoglycoside moieties are positioned in the minor groove. The aminoglycoside, which is attached to spirocyclic ring, aligns along the A20T21G22 sequence of the nonbulged strand, while the dihydronaphthalenone, which is restrained by the spirocyclic structure, is positioned near the G7-T8-T9 bulge site. The aminoglycoside is closely aligned with the dihydronaphthalenone, preventing its intercalation into the bulge site. In the complex, the unpaired purine (G7) is intrahelical and stacks with the intercalating moiety of DDI, whereas the unpaired pyrimidine (T8) is extrahelical. The structure of the complex formed by binding of the synthetic agent to the two-base bulged DNA reveals a binding mode that differs in important details from that of the natural product, explaining the different binding specificity for the bulge sites of DNA. The structure of the DDI-bulged DNA complex provides insight into the structure-binding affinity relationship, providing a rational basis for the design of specific, high-affinity probes of the role of bulged nucleic acid structures in various biological processes.