Imidazole-imidazole pair as a minor groove recognition motif for T:G mismatched base pairs

The T:G mismatched base pair is associated with many genetic mutations. Understanding its biological consequences may be aided by studying the structural perturbation of DNA caused by a T:G base pair and by specific probing of the mismatch using small molecular ligands. We have shown previously that AR-1-144, a tri-imidazole (Im-lm-lm) minor groove binder, recognizes the sequence CCGG. NMR structural analysis of the symmetric 2:1 complex of AR-1-144 and GAAC-CGGTTC revealed that each AR-1-144 binds to four base pairs with the guanine N2 amino group forming a bifurcated hydrogen bond to a side-by-side lm/lm pair. We predicted that the free G-N2 amino group in a T:G wobble base pair can form two individual hydrogen bonds to a side-by-side lm/lm pair. Thus an lm/lm pair may be a good recognition motif for a T:G base pair in DNA. Cooperative and tight binding of an AR-1-144 homodimer to GAACTGGTTC permits a detailed structural analysis by 2D NOE NMR refinement and the refined structure confirms our prediction. Surprisingly, AR-1-144 does not bind to GAATCG-GTTC. We further show that both the lm-lm-lm/lm-Py-lm heterodimer and the lm-lm-lm/lm-lm-lm homodimer bind strongly to the CACGGGTC + GACICGTG duplex. These results together suggest that an lm/lm pair can specifically recognize a single T:G mismatch. Our results may be useful in future design of molecules (e.g. linked dimers) that can recognize a single T:G mismatch with specificity.