Year

2017

Degree Name

Doctor of Philosophy

Department

School of Chemistry

Abstract

Human telomeric DNA is capable of forming four-stranded helical structures known as G-quadruplexes (qDNA), possible substrates of telomerase. Ligand-based stabilisation of telomeric qDNA is a possible method for probing telomerase activity. A small library of berberine derivatives was tested for binding selectivity for different qDNA conformations over double-stranded DNA (dsDNA). The qDNA sequence, 22AG (d[AGGG(TTAGGG)3]), was subjected to different folding conditions to form a range of qDNA conformations. A tetramolecular qDNA structure, formed from four separate strands of 7GGT (d(TTAGGGT)), was also characterised. A dsDNA sequence, D1, (GGAAGGTCCAGAGAGG) (single-strand shown), was used to compare the binding selectivities of the berberine derivatives. Circular dichroism (CD) spectroscopy was used to determine the strand orientation (parallel, antiparallel etc.) and thermal stability of the nucleic acid structures. Intramolecular parallel, mixed hybrid/antiparallel Gquadruplexes, and tetramolecular qDNA were formed in NH4+-containing solutions, allowing the DNA strand and ligand binding stoichiometry for these qDNA structures to be determined using electrospray ionisation mass spectrometry (ESI-MS). Ion mobility ESI-MS was also used to confirm the presence of tetramolecular qDNA. Fluorescence resonance energy transfer (FRET) melting assays were used to determine the stability of antiparallel qDNA and hybrid qDNA formed in Na+- and K+-containing solutions, respectively. Docking and molecular dynamics (MD) simulations were completed for some of the ligand-qDNA complexes to interrogate possible modes of binding.

This thesis is unavailable until Friday, January 04, 2019

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