The covalent and non-covalent interactions between DNA and external ligands and between DNA and itself are critical for cellular function. An increased knowledge of these interactions can be used for the development of disease-fighting agents, specifically anti-cancer drugs with improved sensitivity and specificity for tumor cells. Electrospray ionization mass spectrometry (ESI-MS) is useful in the screening and characterization of the interactions involving nucleic acids given the speed and small sample sizes that can be analyzed. In this dissertation, ESI-MS is used to characterize covalent and non-covalent interactions involving DNA to assist in determining how these interactions can lead to better therapeutics.
The non-covalent binding of ligands to quadruplex oligonucleotides is discussed first. Pyrrole inosine ligands, which bind to guanine bases, were found to interact with both quadruplexes and with guanine rich oligonucleotides without a quadruplex structure. While those interactions were specific with guanine, novel platinum complexes were found to form specific interactions with quadruplex structures themselves as the size of the ligands matched the size of a guanine quartet. This allowed the ligands to end-stack with quadruplexes with large thymine-rich loops between guanine-rich regions.
The non-covalent and covalent interactions between ligands and other DNA structures were also studied. The non-covalent binding of anthracycline ligands to mismatched DNA hairpins was probed. The analysis of solutions of approximately equimolar ligand and oligonucleotide indicated preferential binding to the mismatched sequences. Diazirdinyl benzoquinone crosslinkers, including the clinically studied RH1 and an analogue of RH1, were reacted with a variety of duplex oligonucleotides. The complexes were observed by LC-MS and dissociated using both CID and IRMPD to determine the sites of crosslinking. It was determined that both ligands could form interstrand crosslinks in DNA with 5’-GNC or 5’-GNNC sequences. The RH1 analogue, with a bulky phenyl group, formed fewer crosslinks than RH1.
In addition to studying DNA/ligand interactions, the interactions between oligonucleotides were also probed. Oligonucleotides containing non-standard isoguanine repeats were annealed in the presence of various cations to determine how those cations would affect the resulting secondary structures. In most cases, isoguanine containing strands formed pentaplexes rather than quadruplexes, which were observed for strands containing guanine bases. / text
| Identifer | oai:union.ndltd.org:UTEXAS/oai:repositories.lib.utexas.edu:2152/ETD-UT-2010-05-762 |
| Date | 01 September 2010 |
| Creators | Pierce, Sarah Elizabeth |
| Source Sets | University of Texas |
| Language | English |
| Detected Language | English |
| Type | thesis |
| Format | application/pdf |
Page generated in 0.0021 seconds