This research plan represents a first step toward the design of artificial DNA-interactive agents based on protein motifs that closely resemble known antitumor drugs. These studies seek, in the long term, to develop molecules with pharmacological properties based on the hypothesis that agents designed to imitate DNA binding proteins may exhibit a targeted bioactivity with diminished cytotoxicity in comparison to conventional natural products that associate with DNA. The following proposal will investigate synthetic analogues of a tandem beta-turn motif found in RNA polymerase II (Tyr-Ser-Pro-Thr-Ser-Pro-Ser-Tyr) that: 1) closely resembles antitumor agents of the quinoxaline class (i.e., triostins and echinomycin); 2) binds to DNA in a fashion similar to bis-intercalating drugs; and 3) exhibits a selectivity for AT-rich DNA regions. Specifically, the proposed studies will attempt to increase the structural integrity and DNA binding affinity of this naturally-occurring motif through strategic substitutions of natural and unnatural amino acids and peptidomimetics that are known to support a type II beta-turn conformation. The modifications to be employed have been chosen to minimally perturb the peptide nature of these structures while also representing a first step toward the development of agents with drug-like stability and protease resistance. The structures of these redesigned motifs will be investigated using 2D NMR, circular dichroism (CD) spectroscopy, and molecular modeling. In parallel, investigations of DNA sequence-selectivity through hydroxyl radical footprinting techniques and evaluations of their mechanism(s) of nucleic acid binding will be carried out to fully develop a model of beta-turn-DNA interaction(s). These studies are particularly interested in correlating motif structure and conformational rigidity to DNA selectivity and helical distortion. This direction represents a novel approach to the rational design of agents that bind to DNA; while most studies in this area have been aimed at redesigning and understanding natural products that interact with DNA, these studies will harness and improve upon the designs utilized by DNA binding proteins.
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