SEQUENCE-SPECIFIC RECOGNITION OF DNA BY A DIMER MOTIF

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As part of a drug discovery group to develop anti-opportunistic infection agents that are targeted to DNA-protein complexes, Dr. Wilson has developed unfused aromatic dications that bind in the DNA minor groove. Two of these compounds are now entering clinical trials. An additional exciting discovery from the minor-groove research is that one of the new compounds, a diamidine with a phenyl-furan-benzimidazole ring system (DB293), binds to mixed AT/GC sequences in DNA as a dimer. Initial NMR, DNAse I footprinting, and surface plasmon resonance results clearly support the hypothesis that DB293 binds in the minor groove at specific GC containing sequences of DNA in a highly cooperative manner as a stacked dimer. Previous studies have suggested that such complexes are not possible with dications and neither of the symmetric analogs of DB293 bind significantly to GC sequences. This proposal is built around the hypothesis that DB293 recognizes both strands of DNA and provides a new paradigm for design of compounds for recognition of specific DNA sequences. Such a recognition motif would require reevaluation of ideas on the limits for small molecule-DNA recognition. The general goal of the proposed research is to fully characterize the dimer-DNA binding motif and to develop models that will allow us to extend the mode to additional DNA sequences. A complementary approach utilizing biophysical and synthetic methods will be used to thoroughly characterize the dimer recognition mechanism. Information on the initial dimer complexes will then be used to design, synthesize and characterize the DNA interaction of new compounds with modified and/or extended dimer-DNA sequence recognition capability. Four specific aims will allow Dr. Wilson to achieve these goals: Aim 1: Determine the structural details of the DB293-dimer DNA complex by 2D NMR and x-ray methods if crystals can be obtained. Aim 2: Prepare analogs of DB293 and characterize their DNA interactions to determine what features of the molecular structure are essential for formation of the dimer motif. Aim 3: Modify the DB293-DNA dimer recognition sequence to determine what effect base pair changes have on dimer recognition and affinity. Aim 4: Use all of this information to define the dimer recognition rules for DB293.