The work in this dissertation has two main focuses: (1) to develop sensors based on a quadruplex-forming oligonucleotide scaffold for the sensing of specific sequences, (2) to develop an indicator displacement assay for the high-throughput determination of host-guest binding capable of easy discrimination between strong and weak-binding species. Chapter 1 serves to provide a brief introduction to some shared background for both projects, through introducing basic tenants of aqueous supramolecular chemistry and nucleic acid chemistry, in addition to some general lessons in system design that can be learned from the study of biology. Chapter 2 describes the design of a sensor based on a naphthalene mono-imide (NMI) scaffold, which due to an intramolecular charge-transfer from the naphthalene to a conjugated bipyridine has a low native fluorescence emission. When exposed to curcubit[7]uril (CB[7]) a macromolecular host able to encapsulate the bipyridine unit, the intramolecular charge-transfer is interrupted, resulting in a significant increase in fluorescence quantum yield (by over an order of magnitude). This enhancement is reversible, with competing binders for CB[7] causing a return to the unbound state with quenched emission. Importantly, this sensor exhibits robust activity with no significant variance in properties throughout the range of 5-10 pH, and is amenable to secondary functionalization for surface attachment without loss of activity. A facile microplate assay was developed on the surface-bound sensor, and a proof of concept study was shown by testing for binding against a library of therapeutically relevant drug classes, resulting in the discovery of three novel guests for CB[7] possessing strong to moderate binding affinities. Chapter 3 discusses the development of an oligonucleotide sensor called a quadruplex molecular beacon (QMB) that is able to transition between a closed, intramolecular quadruplex state and an open, intermolecular duplex state on the sequential application of two stimuli in the form of specific oligonucleotide sequences. The chapter initially focuses on optimizing strand exchange mechanisms that allow for the sequential stimuli to open (and close) the system, finding that the combined use of a short toehold sequence with a targeted base-pair mismatch leads to efficient hybridization and displacement with sequential stimuli. The remaining part of the chapter examines the use of this strand exchange mechanism to drive a fully constituted QMB through sequential opening and closing in response to sequential stimuli, with a corresponding fluorescence signal. / Cooper Battle
| Identifer | oai:union.ndltd.org:TULANE/oai:http://digitallibrary.tulane.edu/:tulane_51492 |
| Date | January 2016 |
| Contributors | Battle, Cooper H. (author), Jayawickramarajah, Janarthanan (Thesis advisor), School of Science & Engineering Chemistry (Degree granting institution) |
| Source Sets | Tulane University |
| Detected Language | English |
| Type | Text |
| Format | electronic |
| Rights | Embargo |
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