DEFINING GC-SPECIFICITY IN THE MINOR-GROOVE - SIDE-BY-SIDE BINDING OFTHE DI-IMIDAZOLE LEXITROPSIN TO C-A-T-G-G-C-C-A-T-G

Background: Polyamide drugs, such as netropsin, distamycin and their l exitropsin derivatives, can be inserted into a narrow B-DNA minor groo ve to form 1:1 complexes that can distinguish AT base pairs from GC, b ut cannot detect end-for-end base-pair reversals such as TA for AT. In contrast, 2:1 side-by-side polyamide drug complexes potentially are c apable of such discrimination. Imidazole (Im) and pyrrole (Py) rings s ide-by-side read a GC base pair with the Im ring recognizing the guani ne side, But the reason for this specific G-Im association is unclear because the guanine NH2 group sits in the center of the groove. A 2:1 drug:DNA complex that presents Im at both ends of a GC base pair shoul d help unscramble the issue of imidazole reading specificity. Results: We have determined the crystal structure of a 2:1 complex of a diimid azole lexitropsin (DlM), an analogue of distamycin, and a DNA decamer with the sequence C-A-T-G-G-CC-A-T-G. The two DIM molecules sit antipa rallel to one another in a broad minor groove, with their cationic tai ls widely separated. Im rings of one drug molecule stack against amide groups of the other. DIM1 rests against nucleotides C(7)A(8)T(9)G(10) of strand 1 of the helix, whereas DIM2 rests against G(14)G(15)C(16)C (17) on strand 2. All DIM amide nitrogens donate hydrogen bonds to N a nd O atoms on the floor of the DNA groove and, in addition, the two Im rings on DIM2 accept hydrogen bonds from guanine N2 amines, thereby p roviding specific reading. The guanine N2 amine can bond to Im on its own side of the groove, but not on the cytosine side, because of limit s on close approach of the two Im rings and the geometry of sp(2) hybr idization about the amide nitrogen. Conclusions: Im and Py rings disti nguish AT from GC base pairs because of steric factors involving the b ulk of the guanine amine, and the ability of Im to form a hydrogen bon d with the amine. Side-by-side Im and Py rings differentiate GC from C G base pairs because of tight steric contacts and sp(2) hybridization at the amine nitrogen atom, with the favored conformations being G/lm, Py/C and C/Py,Im/G. Discrimination between AT and TA base pairs may be possible using bulkier rings, such as thiazole to select the A end of the base pair.