PEPTIDE DNA HYBRIDS

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We are interested in determining high resolution structures of peptide-DNA hybrids and using these structures to search for new therapeutic agents that bind to DNA or RNA targets. In particular, peptide sequences that bind tightly to single-stranded oligonucleotides, and thus protect them against nuclease degradation are being sought. Based on the results of the in vitro selection experiments, two peptide-DNA sequences were selected. Two-dimensional spectra (NOESY, TOCSY) of the first sequence (KWKGSGCTTTGAC) were recorded and despite optimization of salt concentration, pH, and temperature, the spectra did not provide sufficient constraints to solve the three-dimensional structure of the highly flexible intramolecular complex. The second sequence (WTG-CGCAC) had been selected from partially self-complementary sequences, where interactions between amino acid residues and the single-stranded region are favored by the template effect of the duplex. NOESY spectra of the chosen hybrid molecule at 40, 80, 160, 320, and 500 ms mixing time were recorded, together with a TOCSY and a ROESY experiment. These spectra were found to contain a rich collection of well resolved cross peaks, allowing us to assign many of the protons in the hybrid. The preliminary molecular dynamics work performed so far with the "simulated annealing" routine of X-PLOR led to a very first set of unrefined structures. Based on these, we are confident that we will be able to solve the structure of this complex.