Includes abstract. / Includes bibliographical references (leaves 101-116). / Oxidoreductases are interesting enzymes with potential applications in a number of different industries such as the textile, food and feed, chemical and biomedical industries. Oxidoreductases require the use of co-factors. These small molecules are relatively expensive and are required in stoichiometric amounts for their enzymatic reaction; this negatively impacts the economic viability of their potential applications. Several methods have been developed to counteract this problem, the most preferred of which is the enzymatic co-factor recycling method. A few methods for the co-immobilisation of enzymes and co-factors have been developed. These systems are of interest as they offer the advantages of recycling the enzymes together with the co-factor, thereby enabling re-use. The immobilisation of enzymes also provides a platform for improving their stability, activity, specificity and selectivity. Since glucose dehydrogenase (GDH) and NADH oxidase, are industrially relevant co-factor recycling enzymes for NAD(P)H and NAD+ respectively, characterisation of their immobilisation is of interest. The current work describes the use of the proprietary particle technology, termed ReSyn™, for the construction of a self-contained co-factor recycling system. The research included the optimisation of immobilisation for the individual enzymes, followed by the co-immobilisation with subsequent co-factor entrapment. The immobilised enzymes displayed improved thermal and pH stability compared to the non-immobilised enzymes. Immobilised GDH also displayed increased activity over the acidic range when compared to free GDH. The system was shown to be capable of recycling NADH/NAD+ up to at least 142 times with a specific activity of 10.18 U.mg¯1. The system was recovered and recycled with a 77% activity efficiency indicating recovery of the system and reusability. Preparation of a functional self-contained co-factor recycling system was demonstrated consisting of the biological components NADH oxidase and glucose dehydrogenase, immobilised on a polyethylenimine support with entrapped cofactor. This serves as proof-of-principle for the construction of derivative systems that could be used for the development of applications such as efficient biosynthesis, novel biosensors, diagnostic and therapeutic systems.

Includes bibliographical references (leaves 114-128). / In this study I explored the possibility of making HIV-1 subtype C Pr55gag-based chimaeric virus-like particles (VLPs) as a boost to the HIV-IC multigene DNA vaccine pTHr.grttnC, which encodes a modified Gag-RT-Tat-Nef fusion protein (GRTTN). Furthermore, an attempt was made to produce VLP analogues to the HIV-IA polyepitope DNA vaccine pTHr.HIVA. A range of in-frame fusions with the C-termini of myristylation-competent p6-truncated Gag and native Pr55gag were made to test how the length of polypeptide and its sequence might affect VLP formation and structure.

Includes bibliographical references (leaves 101-122). / Aquatic species,notably fish, can reveal details of drainage evolution, especially where their evolutionary relationships can be reconstructed using phylogeographic methods. In this study, the mitochondrial DNA sequence diversity of the characiform genus, Hydrocynus, which is widespread across tropical Africa, is reconstructed using a phylogenetic framework and divergences are dated using the cytochrome b (cyt) region.

Includes abstract. / Includes bibliographical references (leaves 153-175). / Cervical cancer is primarily caused by infection with Human papillomavirus (HPV) and is a global concern, particularly in developing countries which contain ~80% of the cervical cancer burden. Current HPV L1 major capsid protein virus-like particle (VLP)-based vaccines are effective in the type-specific prevention of infection and associated disease. However, the high cost of the vaccines has limited their widespread application, and cytological screening programmes are still required to detect malignant lesions associated with the non-vaccine types, particularly in HIV-infected populations.

All of the samples used for actinobacterial isolation were subjected to a culture-independent (metagenomic) study. The results provided an explanation for why no actinomycetes were found in the aquatic samples, as all of the sequenced clones were shown to be most closely related to uncultured bacteria. In the terrestrial sample, a total of 120 clones were obtained and all were sent for sequencing.

Includes bibliographical references (leaves 96-103). / Proteins mediate bio-chemical processes in cellular organisms by functioning as structural components, signaling molecules, enzymes, transcription factors and receptors, through complex interactions that they make with one another. The study of protein-protein interactions (PPI) within an organism is thus a vital step towards understanding the cellular life process of an organism. PPIs have been studied by specially designed experiments to identify pairs of proteins that are suspected to interact. Experimental approaches are quite expensive, yet still there are cases of wrongly identified protein-protein interactions. In this study, we predict PPIs computationally by exploiting known properties of proteins and PPIs and the requirement that proteins be present in the same cell compartment at the same time in order to interact (subcellular localization and gene expression).

Bibliography: leaves 119-150. / Although roots play an integral role in the sensing and amelioration of environmenatl stresses, there are no reports that specifically detail their involvement in the desiccation tolerance mechanisms of resurrection plants. Very lttle is known about even the genral anatomy and physiology of the roots of plants such as X. humilis, and almost nothing about the molecular responses that confer their ability to survive desiccation. This report details foundational studies of the functional anatomy and large-scale molecular responses of the roots during dehydrtion and rehydration.

Bibliography: leaves 108-120. / In this study, we have developed an assay to investigate of the binding mode of yeast linker histone Hholp to nucleosomes, using magnetic bead pull-down assays. To understand the structural and regulatory role of, Hholp, the linker histone in Saccharomyces cerevisiae, and that of higher eukaryotes (Thoma et al., 1979), it is essential to determine the manner in which these histones associate with chromatin. It was previously proposed that Hh01p may bind to two nucleosomes simultaneously, where globular domain one and globular domain two bind to the adjacent nucleosomes.

Includes bibliographical references. / Our group has recently found that the linker histone Hh01 p of Saccharomyces cerevisiae exhibited a significant increase in binding to chromatin during stationary phase. Because of the role of H1 in gene expression and chromatin compaction, it is essential to understand the mechanism behind this change in binding behaviour for a complete mechanistic description of gene regulation. We postulated that the phosphorylation of serine or threonine residues decrease the affinity of H1 for DNA, resulting in the dissociation of H1 from chromatin in exponential phase. We investigated this possible change in the phosphorylation state of Hh01 p in yeast cells in exponential phase and in stationary phase by immunoprecipitation of Hh01 p, followed by western analysis using antiphosphoserine and anti-phosphothreonine antibodies.

Includes bibliographical references. / The XvPSap1 promoter derived from Xerophyta viscosa has been demonstrated to be stress-inducible during dehydration in transgenic Nicotiana tabacum, black Mexican sweetcorn cells and Zea mays. To improve this promoter, for future applications in crop biotechnology, four shortened promoters, XvSap1D, E, F and G were generated by mutagenesis. The generated promoters had circa 50% reduction in size and contained the 5' proximal and 3' distal regions of the XvPsap1 promoter with the internal region removed. The shortened promoters displayed no significant sequence homology to any other known plant promoter, besides XvPSap1. In addition to the shortened promoters, a newly discovered full length XvPSap2 promoter, showing a 56.41% homology with XvPSap 1 was also assessed in this study.