Recent advances in in vitro selection and evolution techniques have identified novel DNA secondary folds selected from random DNA libraries that target immobilized ligands with high affinity and specificity. One component of this application focuses on the structural characterization of ligand-DNA aptamer complexes and DNA metalloenzymes at high resolution through a combined NMR-molecular dynamics approach. These efforts will be extended to the measurement of imino proton hydrogen exchange and base pair opening kinetics on these complexes. Preliminary NMR data are presented on DNA aptamers complexed to AMP, L-argininamide and D-vasopressin, and experiments are outlined to structurally characterize a transition state distorted porphyrin-DNA aptamer complex and a small RNA-cleaving DNA metalloenzyme. The goals of these projects are to identify and to characterize novel three dimensional folds and to understand the molecular basis for the striking selectivity associated with target recognition. The second component of the application addresses NMR based structural and hydrogen exchange characteristics of antitumor drug-DNA complexes. The emphasis in this competing renewal are on antitumor drugs that target the N7-position of guanine through either coordination or covalent binding. These range from cis- and trans-platin that form intrastrand and interstrand cross-links, to mitomycin C monoadducts that form replication blocks, to saccharide containing drugs that target the floor and walls of the DNA helical grooves. The emphasis here is to identify the principles of stacking, hydrogen bonding and hydrophobic interactions that contribute to the specificity and affinity associated with molecular recognition. In addition, a program is proposed to structurally characterize DNA complexes of enediyne based chimeric hybrid drugs prepared in-house in the Samuel Danishefsky laboratory, that mix and match sequence specific and warhead modules in order to generate novel antitumor drugs that target DNA with unique sequence and damage specificities.
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