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A double competition dialysis assay for the analysis of the distribution of optoelectronically active components over nucleic acid structures

Albalawi, Karma 2018. A double competition dialysis assay for the analysis of the distribution of optoelectronically active components over nucleic acid structures. PhD Thesis, Cardiff University.
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This thesis presents DNA binding studies and our work to develop a double competition dialysis assay. Chapter 1 describes DNA structure, including duplex, triplex and quadruplex structures, and functioning in storing the genetic code. This Chapter also presents an overview of the interactions of small molecules with nucleic acids structures. Moreover, the chapter describes the techniques that have been used for our DNA-binding studies, viz, UV - visible spectroscopy, circular dichroism spectroscopy and isothermal titration calorimetry. The chapter also describes potential applictions of small molecule DNA binders. Finally, we describe the competition dialysis in this chapter. Chapter 2 describes the determination of extinction coefficients for selected optoelectronically active π-conjugated molecules in aqueous buffers. Furthermore, we established the light sensitivity of the compounds. In addition, the chapter describes the binding studies of nucleic acid binders from a library of available ligands using UV-visible, circular dichroism, and isothermal titration calorimetry. Chapter 3 describes the development of a custom competition dialysis device. We test this device to determine affinity and selectivity of ligands for nucleic acids structures. We analysed the affinity and selectivity of a single ligand for FS-DNA, specific duplex sequences (dAdT)12●(dAdT)12 and (dGdC)12●(dGdC)12, and different quadruplex structures such as cmyc, 22AG and EAD2. The data agree with the results from UV-vis titrations. In Chapter 4 we explore how double competition dialysis allows screening of two ligands against an array of nucleic acids structures. Several compounds were tested showing that our assay deals reasonably well with fading unless the latter progresses to the extent when absorbance is too low to measure reliably. Although we have identified compounds with promising affinity profiles, even in the presence of a second binder we are yet to identify binders with an orthogonal selectivity profile. In Chapter 5 we present general conclusions and suggestions for future work.

Item Type: Thesis (PhD)
Date Type: Completion
Status: Unpublished
Schools: Chemistry
Subjects: Q Science > QD Chemistry
Date of First Compliant Deposit: 12 February 2019
Last Modified: 03 Aug 2022 01:48

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