Allostery is a form of signalling within biomolecules such that ligand binding to a protein affects its activity at a second site. Allostery was described by early models to be driven by structural changes in the protein. However, more recently there has been increasing evidence that dynamics can contribute to or even drive allostery. The protein studied in this thesis, the Catabolite Activator Protein (CAP), is an allosteric protein homodimer that has been shown to exhibit negatively cooperative binding of the ligand cyclic Adenosine Monophosphate (cAMP) to each of its monomers. Interestingly, CAP is a protein whose allostery is believed to be driven by dynamics rather than a conformational change. In this thesis, a number of coarse grained models are employed to investigate this dynamic allostery in CAP. One family of models, termed Super Coarse Grained (SCG) models explore the global properties of the dynamics of the CAP dimer that cause it to exhibit negatively cooperative allostery. It is shown through these models that changes in protein interactions can provide a basis for changing cooperativity. A second family of coarse grained models called Elastic Network Models (ENM) are studied. These are used to show that adjusting the interactions between specific residues can affect cooperative binding of cAMP to CAP. A number of atomistic approaches are also used to study the cAMP-CAP system, including Molecular Dynamics (MD) and Normal Mode Analysis (NMA). The efficacy of using such approaches for studying the thermodynamics of the allostery in CAP is investigated. The motion observed within the protein is also studied closely to identify potential allosteric pathways. X-ray crystallography and Isothermal Titration Calorimetry (ITC) are finally used to investigate how accurately computational methods can describe the cooperative binding of cAMP to CAP. They are also used to try and determine whether the allostery in CAP can be manipulated experimentally without any observed changes to its structure.

Dilute solutions of high molecular weight (HMW) polymers can reduce friction experienced by a fluid in turbulent pipe flow, greatly decreasing energy required for transfer of the solution. These polymers are known as drag reducing agents (DRA). Polyacrylamide (PAM), synthesised using free-radical inverse-emulsion polymerisation, is most commonly used in commercial aqueous applications. Restrictions on the acrylamide monomer have recently been imposed due to its carcinogenicity, furthermore, the use of inverse-emulsions, containing oil and surfactant, has a negative impact on the environment. The susceptibility of HMW polymers to mechanical degradation in turbulent flow quickly decreases their drag reducing efficiency (DRE), a major problem for current systems. The aim of this project was to produce an effective water soluble polymer DRA with the following properties; 1) acrylamide free; 2) environmentally friendly; 3) oil/surfactant free; 4) mechanically stable; 5) economically viable. It was proposed that by the synthesis of HMW, acrylamide free, star polymers using Cu(0)-mediated polymerisation, effective water soluble drag reducing polymers with enhanced mechanical stability could be produced. An aqueous polymerisation method at ambient temperature would greatly reduce the environmental impact of the process. In Chapter 1, a general background of the drag reduction phenomenon, potential mechanisms of action and key properties for effective drag reducing polymers is given. This chapter also introduces branched polymers and controlled radical polymerisation methods; in particular, Cu(0)-mediated techniques. Chapter 2 focusses on the synthesis of water-soluble, poly(ethylene glycol) (PEG) containing macro-initiators (I4-S, I4-T and I2-S) for use in Cu(0)-mediated polymerisation reactions. By coupling a branching unit containing two potential initiation sites to each end of a PEG chain, multi-functional initiators for the synthesis of star polymers were produced and fully characterised using 1H and 13C NMR spectroscopy as well as MALDI-ToF mass spectrometry. In Chapter 3, the polymerisation of tert-butyl acrylate (tBA) is conducted in DMSO utilising a several initiators; 4,4’-oxybis(3,3-bis(2-bromopropionate)butane (4AE), I4-S, I4-T and I2-S, and a simple catalyst system (Cu(0)/TREN). The reactions proceeded as a self-generating bi-phasic system due to the insolubility of PtBA in the solvent. A model initiator, methyl 2-bromopropionate (MBP), was also used to investigate the polymerisation of tBA further by introducing several changes in reaction conditions. The initiators were used to prepare polymer samples for drag reduction testing. Chapter 4 describes the aqueous Cu(0)-mediated polymerisation of sodium acrylate (NaA) using the I4-S and I4-T macro-initiators. Kinetics study of the reaction using I4-T demonstrated the polymerisation proceeded via a free-radical mechanism. Although complete control over the reaction was not observed, a branched polymer was synthesised by the incorporation of the multi-functional macro-initiator in to the final product and HMW samples were generated for drag reduction testing. The drag reducing properties of the PtBA and PNaA samples are tested in Chapter 5 using a pipe flow test rig. The PtBA samples were first hydrolysed using trifluoroacetic acid (TFA) to provide water soluble poly(acrylic acid) (PAA). For comparison, several commercially available HMW PAM (Praestol, PAM-6M), poly(ethylene oxide) (PEO-8M) and PAA (PAA-1M) samples were also measured. The results demonstrated that the branched PAA/PNaA samples were effective as DRAs. Furthermore, by cycling the polymer solution through the test rig the mechanical stability of the polymer samples was investigated. An increased resistance to mechanical degradation was observed for the star polymers when compared to linear analogues. In Chapter 6 general conclusions and future perspectives for the work are discussed.

The thesis deals with enzymatic catalysis in bicontinuous microemulsions, which consist of a dynamic network of oil and water domains separated by a monolayer of surfactant molecules, i.e. the interfacial layer. Hence, a microemulsion with the composition buffer – n-octane – nonionic surfactant was tested as a reaction medium for enzyme-catalysed reactions with the emphasis on the conversion of hydrophobic substrates, which are difficult to convert in aqueous buffer solutions. The first part of the thesis focuses on the activity of the lipase B from Candida antarctica (CalB) in bicontinuous microemulsions. First, the optimum reaction conditions determined by temperature, pH and ionic strength were evaluated. Second, it was found that CalB concentrations which showed fast adsorption at an oil-water interface also displayed fast reaction rates. Additionally, no saturation was found for substrate concentrations up to 40 mM of p-nitrophenyl laurate, which according to Michaelis-Menten suggests a Km >> 40 mM. Third, the composition of the interfacial layer had a distinct influence on CalB activity, e.g. the presence of sugar surfactants (b-C10G1) or phospholipids (DOPC) enhanced or decreased CalB activity, respectively. The second part of the thesis describes the activity of the squalene-hopene cyclase from Alicyclobacillus acidocaldarius (Aac SHC) converting its natural substrate squalene in bicontinuous microemulsions. The Aac SHC activity studies revealed a linear dependence on enzyme concentration and a hyperbolic curve for the substrate concentration, with a saturation of Aac SHC at substrate concentrations above 20 mM. The composition of the interfacial layer was found to have neither a significant influence on the activity nor on the conformation of Aac SHC. In summary, good turnover rates were achieved for interfacially-active enzymes (CalB) due to enhanced enzyme-substrate contact at the interfacial layer. For water-soluble enzymes (Aac SHC), a distinctly enhanced selectivity was discovered, although no faster reaction rate was found. The main difference in the catalytic turnover was explained by the adsorption of CalB at the interfacial layer, whereas Aac SHC stays in the aqueous phase of the microemulsion. To conclude, bicontinuous microemulsions were suitable for enzymatic catalysis and are thus interesting in terms of reaction medium engineering to optimise biocatalytic processes.

Foot-and-mouth disease virus (FMDV) causes a highly infectious disease of cloven-hoofed livestock with economically devastating consequences. The political and technical problems associated with the use of FMDV vaccines make drug-based disease control an attractive alternative. The FMDV genome is translated as a single polypeptide precursor that is cleaved into functional proteins by virally-encoded proteases. Ten of the thirteen cleavages are performed by the highly conserved 3C protease (3Cpro), making this enzyme an obvious target for anti-viral drugs. This thesis details an attempt to develop 3C protease inhibitor with efficacy against FMDV. This includes both rational drug design based on the peptide substrate of the protease, and also the high throughput screening of small molecules.

Nanocrystalline inorganic semiconductors have emerged as attractive alternatives to molecular dyes as photosensitisers for mesoporous metal oxide-based semiconductor-sensitised solar cells (SSSCs). In this configuration, Sb2S3 has shown particular promise, yielding impressive power conversion efficiencies. At the operational heart of these devices, there are a series of interfacial electron transfer reactions that act to separate photogenerated charges. The delicate balance between charge separation and recombination plays an important part in defining device efficiency, but the mechanism by which charge separation occurs in Sb2S3-based systems has been relatively poorly understood. In the first part of this work, transient absorption spectroscopy is used to probe interfacial electron and hole transfer in mesoporous TiO2/Sb2S3-based photovoltaic assemblies. The reported observations point to the importance of the hole transfer reaction not simply as a means of regenerating the sensitiser, but rather as an integral part of the charge separation process. Inspired by these results, a novel device architecture and processing route is developed in which the structural, optical and electrical properties of the sensitised TiO2 film are encompassed in a single component; namely a mesoporous Sb2S3 photoanode. The resulting high surface area is shown to allow for efficient charge transfer to a polymeric hole acceptor, whilst electron transport through the Sb2S3 layer is demonstrated by the fabrication of functioning photovoltaic devices. In order to consider the role of mesostructure in Sb2S3-based solar cells, devices based on a flat Sb2S3/hole transport material (HTM) interface are developed and compared to SSSC analogues. It is found that although these two classes of device are structurally very different, overall performance is remarkably similar. This highlights the impressive performance of the thin-film assembly, but also draws into question the importance of meso-scale morphological control. Through a combination of transient absorption spectroscopy and transient photovoltage measurements, it is determined that structural limitations to interfacial charge separation can be overcome in these systems by the action of an electric field. Understanding the role of mesostructure is a particularly pertinent challenge in the context of organic lead halide perovskite absorbers, with which very high efficiencies have been achieved in a range of structured and non-structured device architectures. Here, it is shown that the presence of a mesostructured electron acceptor to rapidly quench the perovskite excited state enhances the stability of interfacial charge separation and significantly increases innate tolerance to environmental processing conditions. This work highlights a significant advantage of retaining mesoscale morphological control in the preparation of efficient, low cost and stable hybrid photovoltaics.

Polymer:fullerene blend microstructure has been recognised as a key, but poorly understood, factor in optimising performance of polymer:fullerene based solar cells. This thesis focuses on investigating the impact of material chemical structure upon the blend microstructure, and thereby on device efficiency and stability for polymer:fullerene solar cells. It will be shown in the first results chapter that polymer fluorination can promote phase segregation in polymer:fullerene blend, limiting its charge generation as demonstrated using transient absorption spectroscopy. The promotion in phase segregation is shown to vary from polymer to polymer. With a careful modification of the polymer backbone, the negative impact of polymer fluorination can be avoided, resulting in an improvement in device performance. The following chapter focuses on the impact of polymer side chain on device performance. Linear side chain for PTTV polymers is found to offer a better polymer:fullerene mixing in comparison to branched side chains, potentially due to its better ability to accommodate fullerene molecules. This subsequently addresses limitations of poor of fullerene exciton dissociation efficiency. The next chapter provides new insights into the effects of polymer molecular weight on device performance. A high molecular weight of two DPP-containing polymers is shown to be beneficial to device performance, by offering a higher degree of material mixing in the polymer:fullerene blend. The last results chapter compares two polymers with different crystallinity in terms of their blend microstructure and device performance upon thermal annealing. It is shown that devices employing a crystalline DPP based polymer exhibits a sharp collapse in efficiency for annealing at temperatures beyond 140 °C, which is assigned to the polymers poorer miscibility with fullerene, making it incapable to suppress fullerene clusters formation. In contrast, an amorphous IDT based polymer shows a smaller decrease in efficiency under the same condition, consistent with its greater miscibility with fullenere.

The observation of photocatalytic water splitting on the surface of anatase TiO2 crystals has stimulated many investigations of the underlying processes. Nevertheless, a molecular level understanding of the reaction is not available. Therefore, the work carried out here is focused on improving the understanding of the water splitting mechanism by studying anatase TiO2 surfaces with hybrid-exchange density functional theory calculations. Initially, a simple and systematic methodology for the simulation of constant current scanning tunnelling microscopy (STM) images is developed in order to study TiO2 surfaces. The methodology has to overcome a significant limitation of local basis set calculations: the poor description of the charge density in the vacuum region above the surface. The methodology is tested on various surfaces and the simulated STM images are found to accurately reproduce experimental data. The next step was to re-establish the atomistic structures of the (101), (001) and (100) surfaces. The relative stability of these surfaces is interpreted in terms of the coordination of the surface ions and the geometry surrounding them. Furthermore, the electronic properties are studied. Based on relative positions of the conduction and valence bands of the low-index surfaces studied the (101) and (001) surfaces were identified as the most likely destinations for photo-generated electrons and holes, respectively. The atomistic structure of two surfaces vicinal to the (101) surface - the (514) and (516) surfaces - was established. The surface formation energy of these surfaces is low and the (516) surface contributes significantly (27%) to the surface area of an equilibrium crystallite, which could suggest that this surface is exposed in nanoparticles. The conduction band edge of this surface was computed to be more negative than that of the (101) surface. Thus, if the (516) surface is exposed in nanostructures, it would replace the (101) surface as the most likely destination for photo-generated electrons. Under-coordinated ions at the steps of these vicinal surfaces were identified as preferential adsorption sites and water adsorption was studied at these sites. In general, it was found that water preferentially adsorbs in the molecular adsorption mode on all sites tested, with the exception of the Ti4c on the (514) surface, where the dissociative mode is preferred. Nevertheless, at the (516) step the binding energy of the dissociative adsorption mode is comparable to that of the molecular case. Therefore, if it were possible to preferentially expose the Ti5c-O2c-Ti5c motif from the (516) surface, or even the Ti4c of the (514) surface the reactivity for water photolysis could be enhanced.

Transition metal-free, thermal cyclisation methodology has been demonstrated previously in the Parsons group, with the vision of applying the process to a novel synthesis of the antibiotic Lactonamycin. i. The synthesis of the aglycone of i has been proposed from the pentacyclic intermediate ii, which itself is the hypothesised product of the Parsons-BoardWaters cyclisation of the ene-diyne iii. (Scheme A - see thesis abstract) The phthalide v was synthesised from 2,5-dimethoxybenzaldehyde in 3 steps in moderate yield and was seen as a common starting material for the subsequent routes devised to provide the key aldehyde vi. A further mode of complexity was proposed by attempting to perform the Claisen rearrangement of the alkene vii in cascade with the aforementioned cyclisation to give the amide ii. (Scheme B - see thesis abstract)

p97 is an abundant protein in human cells and is essential for many forms of life. Its putative mechanism of action is the binding of adaptor proteins, which mediate its functions, and subsequent transfer of energy from ATP hydrolysis through the adaptor to substrate proteins. The unfolding or degradation of these substrates is known to regulate a diverse number of processes and the malfunction of p97 in many of these has been linked to a range of diseases. Developing inhibitors for p97-cofactor binding may help uncover further functions, as well as potentially providing a treatment for a number of the diseases in which p97 is implicated. A high-throughput assay based on Förster Resonance Energy Transfer (FRET) was developed, which allowed the high-throughput analysis of candidate compounds. Using this assay, the affinity of p97 for several of its partner proteins was measured. The results were found to be similar to literature values and the assay was measured to have a high Z' factor. Analysis of the hot-spot interactions between p97 and the adaptor protein, p47, enabled the design of novel peptide inhibitors of p97-Cofactor binding. A range of small molecules were also identified through a computer modelling approach known as scaffold hopping. A peptide that was designed to closely mimic the S3/S4 loop within p47 was found to bind to p97 and inhibit its interaction with p47. No inhibition, however, was detected from a second generation of peptides. Two molecules from the virtual ligand screen were found to inhibit p97 binding. Of these, one of the compounds was an unnatural tripeptide, amenable to the rapid synthesis of multiple variations. A second generation of compounds based on this original hit was analysed and another compound was identified with an improved level of inhibition.

No description available.