Electrospray ionization tandem mass spectrometric techniques for the analysis of drug/DNA complexes

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Title: Electrospray ionization tandem mass spectrometric techniques for the analysis of drug/DNA complexes
Author: Mazzitelli, Carolyn Leigh, 1979-
Abstract: Many anticancer and antibacterial therapies are based on the interaction of small molecule drugs with DNA . Increasing interest in the development of DNA -interactive agents has fostered the need for sensitive and versatile analytical techniques that are capable of characterizing the DNA /ligand interactions and are compatible with librarybased screening methods . Electrospray ionization mass spectrometry (ESI -MS ) has emerged as a useful technique for the analysis of non -covalent complexes formed between DNA and small molecules due to its low sample consumption and fast analysis time . The work presented in this dissertation is aimed at exploring , optimizing , and validating ESI -MS methods for characterizing DNA -ligand interactions . ESI -MS is first used to assess the binding of threading bis -intercalators to duplexes containing different sequences to determine high affinity binding sites of the ligands . Preliminary DNAse footprinting experiments identified possible specific binding sites of the ligands and ESI -MS experiments revealed that the ligands bound to DNA duplexes containing the respective specific binding sequences . The metal -mediated binding of benzoxazole ligands with different side chains to duplex DNA is also examined . Cu²⁺ and Ni⁺ were found to promote the most dramatic increase in ligand binding , and ligands exhibiting the most dramatic metal -mediated or metal -enhanced binding were also determined to be the most cytotoxic . The quadruplex DNA binding selectivity of perylene diimides is evaluated by screening the binding of the ligands to quadruplex , duplex and single strand DNA by ESI -MS . Three ligands , one containing basic side chains , one containing anionic sidechains , and one benzannulated compound were determined to be the most -quadruplex selective . The ESI -MS results correlated well with spectroscopic experiments . The relative gas -phase stabilities of different quadruplex DNA structures were investigated using molecular dynamics simulations and ESI -MS . The stabilities from the E[subscript 1 /2] values generally paralleled the RMSD and relative free energies of the quadruplexes based on MD energy analysis . Finally an ESI -MS technique employing the KMnO₄ reaction with DNA to determine conformational changes to the duplex structure upon ligand binding is detailed . Thymines in most intercalator /duplex complexes are more susceptible to oxidation by KMnO₄ than those in duplex DNA . CAD and IRMPD experiments are used to identify the site of oxidation .
URI: http : / /hdl .handle .net /2152 /3156
Date: 2008-08-28

Citation

Electrospray ionization tandem mass spectrometric techniques for the analysis of drug/DNA complexes. Doctoral dissertation, The University of Texas at Austin. Available electronically from http : / /hdl .handle .net /2152 /3156 .

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