Abstract Collagen prolyl 4-hydroxylases catalyze the hydroxylation of proline residues in collagens. The vertebrate enzymes are α2β2 tetramers in which the β subunit is identical to protein disulphide isomerase (PDI). Two isoforms of the catalytic α subunit have been identified in vertebrates, forming type I [α(I)]2β2 and type II [α(II)]2β2 collagen prolyl 4-hydroxylase tetramers. This thesis reports on the cloning and characterization of a third vertebrate α subunit isoform, α(III). The recombinant human α(III) isoform associates with PDI to form an active type III collagen prolyl 4-hydroxylase tetramer, and its Km values for the cosubstrates are very similar to those of the type I and II enzymes, those for a peptide substrate and an inhibitor being found to lie between the two. The α(III) mRNA is expressed in all tissues studied but at much lower levels than the α(I) mRNA. A novel mixed tetramer PHY-1/PHY-2/(PDI-2)2 was found to be the main collagen prolyl 4-hydroxylase form produced in the nematode Caenorhabditis elegans in vivo and in vitro. However, mutant nematodes can compensate for the lack of the mixed tetramer by increasing the assembly of PHY-1/PDI-2 and PHY-2/PDI-2 dimers, these forms also being unique. The catalytic properties of the recombinant mixed tetramer were characterized, and it was shown by the analysis of mutant worms that PHY-1 and PHY-2 represent the only catalytic subunits needed for the hydroxylation of cuticular collagens. The roles of the two catalytic sites in a collagen prolyl 4-hydroxylase tetramer were studied by using the C. elegans mixed tetramer and a hybrid C. elegans PHY-1/human PDI dimer. An increase in the chain length of the peptide substrate led to an identical decrease in the Km values in both enzyme forms. It is thus clear that two catalytic sites are not required for efficient hydroxylation of long peptides, and their low Km values most probably result from more effective binding to the peptide-substrate-binding domain. Inactivation of one catalytic site in the mixed tetramer reduced the activity by more than 50%, indicating that the remaining wild-type subunit cannot function fully independently.

The Arizona Child Acoustic Database consists of longitudinal audio recordings from a group of children over a critical period of growth and development (ages 2-7 years). The goal of this database is to 1) document acoustic changes in speech production that may be related to physical growth 2) inform development of a model of speech production for child talkers. This work was funded by NSF BSC-1145011 awarded to Kate Bunton, Ph.D. and Brad Story, Ph.D, Principal Investigators. This database contains longitudinal audio recordings of 55 American English speaking children between the ages of 2-7 at 3-month intervals. Since children began the study at different ages, some children have fewer recording sessions than others. The database can also be used to provide cross-sectional data for children of a specific age. Please refer to the subject data table for information on specific sessions available here http://arizona.openrepository.com/arizona/handle/10150/316065. All children were recorded using the same protocol; therefore, task numbers are consistent across children and sessions. A calibration tone is included as Record 1 for all sessions. The speech protocol focused on production of English monopthong and diphthong vowels in isolation, sVd, hVd, and monosyllabic real words. In addition, the protocol includes several nonsense vowel-to-vowel transitions. Speakers were prompted either verbally by investigators or by graphical prompts. Details of the protocol with reference to task numbers can be found in the protocol spreadsheet available here http://arizona.openrepository.com/arizona/handle/10150/316065. Details on data recording: All samples were recorded digitally using an AKG SE 300B microphone with a mouth to mic distance of approximately 10 inches. Signals were recorded digitally using a Marantz PMD671, 16 bit PCM (uncompressed) at 44.1KHz. Recordings are made available in .wav format. Individual zip files contain all recordings from a single session.

Malignant melanoma is a growing problem with more and more people in Sweden and the world suffering from this cancer. Malignant melanoma is a disease that when discovered in time can be treated successfully with surgical methods, but the real challenge lies in treating the disease after its spread. Treatment in the past for advanced malignant melanoma has been unsuccessful with no positive effect on overall survival. However, in the last couple of years, new treatment has arrived with focus on priming the immune system to eradicate the tumors. One new drug is the CTLA-4 inhibitor ipilimumab that is given as intravenous infusion. CTLA-4 is a protein located on regulatory T-cells and that is upregulated on activated cytotoxic T-cells. This protein mediates an inhibitory signal that attenuates T-cell-activation.  Treatment with the CTLA-4 inhibitor has been shown to increase overall survival. However, not much is known about how well ipilimumab synergizes with other drugs used for treatment of malignant melanoma.   This is a literature study with the aim to evaluate if ipilimumab is used best as monotherapy or if it is of better use as part of a combination therapy. Search was made in PubMed with the key-words "Ipilimumab", "Ipilimumab treatment", CTLA-4 inhibitor" and "treatment malignant melanoma”. Six articles were chosen and each of these analyzed the effect of ipilimumab alone or combined with other agents against malignant melanoma.   The combination of ipilimumab and the alkylating agent dacarbazine was shown to have a better impact on overall survival compared with monotherapy with dacarbazine, but this combination also showed an increase in serious adverse events. Ipilimumab also showed to work in synergy with both the PD-1-inhibitor nivolumab and the granulocyte macrophage colony-stimulating factor (GM-CSF) sargramostim. Combination with sargramostim was also shown to decrease the amount of serious adverse events. Combination with a gp100 peptide vaccine failed to show any positive effects on overall survival. Also prophylactic treatment with budesonide showed no further gain in overall survival. The effect of ipilimumab was found to have a dose-ranging effect, with higher dose treatment having a better effect but also with more serious adverse events.   The results of this literature study showed that ipilimumab has a better effect with higher dose and that it can work in synergy with other agents such as nivolumab and sargramostim. Results also showed that occurring adverse effects during treatment with ipilimumab may be treated with systemic glucocorticoids that did not affect the tumor-killing ability of ipilimumab. These results should be evaluated in bigger studies and with longer follow up time.

Ces travaux de thèse ont porté sur le développement d’une méthodologie de conversion de chiralité centrale vers axiale pour la formation de 4-arylpyridines atropoisomères, et de son application en synthèse totale. En premier lieu, une méthodologie de synthèse organocatalysée a été optimisée pour la préparation de 1,4-dihydropyridines énantioenrichies et hautement encombrées. Le défi a été, ici, de réussir à trouver le bon compromis entre sélectivité et réactivité afin de générer suffisamment d’encombrement sur la position C4 de la 1,4-dihydropyridine énantioenrichie, et de pouvoir accéder, après conversion du centre stéréogène (C4) en axe de chiralité, à un atropoisomère 4-arylpyridine stable. Une optimisation des conditions opératoires pour l’oxydation de ces 1,4-dihydropyridines énantioenrichies en 4-arylpyridines correspondantes a ensuite été menée et a permis d’atteindre des conversions de chiralité modérées à totales. Sur la base de cette stratégie, la synthèse énantiosélective de la (+)-streptonigrine, produit naturel présentant un motif 4-arylpyridine, a été envisagée selon deux stratégies principales s’appuyant sur des processus organocatalysés. / This work focused on the development of central-to-axial chirality conversion methodology for the synthesis of 4-arylpyridine atropisomers, and its application in total synthesis. In the first place, synthetic methodology was optimised for the synthesis of enantioenriched and hindered 1,4-dihydropyridines. At this point, the challenge was to find the right compromise between selectivity and reactivity to get enantioenriched dihydropyridines with sufficient bulkiness around the C4 position, for formation of stable 4-arylpyridine atropisomers after conversion of the chiral center (C4) to a chiral axis. A detailed screen was performed to find the optimal oxidation conditions leading to moderate to full chirality conversion. Based on this strategy, the total synthesis of (+)-streptonigrin, a natural product containing a 4-arylpyridine framework, was planned following two main pathways using organocatalytic transformations as key steps.

Young people face escalating unemployment rates and insufficient or unsatisfactory job opportunities. Despite this, most of them prefer a career in a formal organisation to a career in entrepreneurship. To more effectively prompt young people to choose entrepreneurship as a career, it is critically important to understand the personal and environmental factors relating to entrepreneurial career choice: the antecedents of entrepreneurial career choice. The core research question underlying this study is: What are the factors relating to entrepreneurial career choice in young people? This study first constructs a conceptual framework of antecedents starting from the extant literature in this field. It then attempts to validate the framework empirically in a sample of South African secondary-school students, laying the foundation for building a theory of antecedents of entrepreneurial career choice. The data collection instrument is a questionnaire compiled mainly on the basis of validated questions available in the literature. The sample is made up of Grade-10, -11 and -12 secondary school learners, aged sixteen, seventeen and eighteen, respectively, selected using a convenience sampling technique. Different statistical tests are employed to examine the relationship between each antecedent factor of the conceptual framework and entrepreneurial career choice. The majority of respondents expressed entrepreneurial-career-choice intentions. The results confirmed a relationship between entrepreneurial career choice and the following antecedents: family support, community support, early childhood experiences, work experience, prior start-up experiences and education. An intricate network of relationships was also found among the antecedents investigated, including some biographical variables. The findings suggest that the current Economic and Management Science (EMS) curriculum implemented in South African secondary schools is achieving its objective of raising students’ entrepreneurial-career-choice intentions. At the same time, however, a number of extra-curricular activities and experiences seem to be related to students’ entrepreneurial-career-choice intentions. This signifies that efforts to foster the choice of an entrepreneurial career among the youth should not be focused entirely on adjusting the school curriculum. Lastly, the number of inter-relationships observed among the antecedents of entrepreneurial career choice signifies that entrepreneurial career choice is a complex phenomenon and that building a theory of antecedents is an arduous undertaking. / Thesis (PhD)--University of Pretoria, 2013. / Business Management / unrestricted

The reporting of the financial results of an organisation is the responsibility of the management of that organisation. However, value may be added to the financial statements by the auditing of such financial statements and by the opinion expressed by the external auditors. Furthermore, there is the expectation on the part of the users of the financial statements that the auditors are also responsible for detecting fraud and, more specifically, financial statement fraud. It was stated in the Association of Certified Fraud Examiners 2010 Report to the Nations that it is the high-level perpetrators who cause the greatest damage to their organisations. The costs arising from financial statement fraud were found to be more than three times higher than the costs arising from fraud committed by lower-level managers and nine times more than the costs involved in employee fraud. The question, thus, arises as to why the auditors would not detect financial statement fraud timeously. The external audit profession has formulated a specific standard which addresses the responsibility of the external auditor as regards the detection of fraud during the audit of financial statements. The aim of this research was to determine the adequacy of the internal auditor standards as regards providing guidance to the internal auditors in terms of detecting financial statement fraud. This research highlighted the lack of guidance in the internal audit standards regarding the responsibility of internal auditors relating to financial statement fraud. In the main, both the directives and the guidance refer to fraud in general but not specifically to financial statement fraud and, thus, the professional internal auditor is forced to seek guidance outside of the internal audit standards as regards the detection of financial statement fraud. / MPhil University of Pretoria, 2014 / Auditing / MPhil / Unrestricted

Elaboration de matériaux métallacalix[4]arènes pour la valorisation des hydrocarbures basée sur un nouveau concept de greffage par voie COMS dont la mise au point par fonctionnalisation de supports silice en faisant réagir l’hydroquinone avec une espèce originale [(SiO)2AliBu.(Et2O)]. La réactivité du complexe W(CtBu)(CH2tBu)3 avec l’hydroquinone supportée [(SiO)2AlO-C6H4-OH.(Et2O)] a permis de générer un nouveau système catalytique bien défini [(SiO)2AlO-C6H4-OW(CtBu)(CH2tBu)2.(Et2O)] entièrement caractérisé. Par cette méthode, l’ancrage du calixarène [[4H]-(OH)3(H)] sur le complexe de surface [(SiO)2AliBu.(Et2O)]SBA-15-(700) a conduit à l’espèce [(SiO)2Al-O-[4H]-(OH)2.(Et2O)]. L’incorporation de complexes organométalliques du groupe IV (Zr) et VI (W) mène à de nouveaux matériaux métallacalix[4]arènes bipodaux. L’élucidation de leurs structures a été appuyée par comparaison avec les données spectroscopiques de leurs homologues solubles ou directement liés à la silice. Ces matériaux, présentant des espaceurs phénoxy, montrent des activités initiales supérieures à celles de leurs homologues supportés sur silice. Avant la préparation des matériaux métallacalix[4]arènes, une série de complexes titana-, zircona- et tantalacalix[4]arenes solubles bi- et tripodaux a été synthétisée et caractérisée à partir de dérivés calix[4]arène présentant des modes de coordination différents pour le métal (podalité, ligands ancillaires, angle OMO). Parallèlement, une relation structure-réactivité dans la polymérisation des oléfines a été établie dans le cas des titanacalix[4]arènes, et pour les tantalacalix[4]arènes, des activations Csp2-H et O-Me intramoléculaires ont été mises en évidence. / Generation of novel metallacalix[4]arenes materials applied to the valorization of hydrocarbons prepared using an unprecendented COMS method by the first functionalization of the silica support by reaction of hydroquinone with the unreported supported species [(SiO)2AliBu.(Et2O)].The reactivity between the complex W(CtBu)(CH2tBu)3 and the hydroquinone species [(SiO)2AlO-C6H4-OH.(Et2O)] allows the access to new well-defined catalytic systems [(SiO)2AlO-C6H4-OW(CtBu)(CH2tBu)2.(Et2O)]. Using this way, the anchoring of the calixarene [[4H]-(OH)3(H)] onto [(SiO)2AliBu.(Et2O)]SBA-15-(700) leads to the surface complex [(SiO)2Al-O-[4H]-(OH)2.(Et2O)].The subsequent incorporation of organometallic complexes from groups IV (Zr) and VI (W) provided novel bipodal metallacalix[4]arenes materials, characterized by IR, solid-state NMR, microanalysis and EXAFS. The clarification of their structures was supported by comparison of the spectroscopic data collected from their soluble models or analogues directly bonded to silica. These new materials, presenting phenoxo linkers display higher initial conversion rates than their analogues directly grafted on silica. Before preparation of the materials, a set of bi and tripodal titana-, zircona- and tantalacalix[4]arenes complexes were synthesized and characterized from calix[4]arenes derivatives ligands, presenting different mode of coordination for the metal (podality, ancillary ligands, OMO bite angle). In parallel, a structure-reactivity relation in the polymerization of ethylene was established in the case of the titanacalix[4]arenes, while Csp2-H and O-Me intramolecular activations were related for the tantalacalix[4]arenes.

Class of 2015 Abstract / Objectives: Two characterized peptide substrates were assayed with human Polo-like kinase 4 to determine phosphorylation activity. A pilot library of Type-II kinase inhibitors designed to fit into the ATP-binding pocket will be screened to determine HsPlk4 inhibition activity, which will help characterize a novel drug compound. Methods: Two peptide substrates of varying concentrations (2 uM, 1 uM, and 0.5 uM) were each combined with serial dilutions of HsPlk4 (1.25 uM, 0.625 uM, 0.313 uM, 0.156 uM, 0.078 uM, and 0.039 uM). EZ Reader detected phosphorylation activity by measuring fluorescence of both substrate and product, which separated at respective time points based on electrophoresis. The subsequent part of the experiment will be to inhibit the kinase activity with molecular inhibitors. Results: The results showed HsPlk4 activity with the modified PLKtide, (5FAM)KKKTPSDSLYDDGLSKK(CONH2). All reactions with the various concentrations of substrate 1 and HsPlk4 showed phosphorylation activity. The reaction started within the first 10 minutes, quickly reaching maximal phosphorylation of substrate. No p-values were calculated due to lack of data. Conclusions: No overall conclusions can be drawn based on the current results. Results showed the reaction reached its saturation point, so methods need to be refined to obtain data within the first 10 minutes. HsPlk4 phosphorylation of PLKtide confirmed the presumption that PLK family is a conserved family of Ser/Thr kinases. There are practical limitations for obtaining good kinetics data depicting enzyme activity, such as having EZ Reader quickly sample the reaction.

The motivation of my research is the massive problem of physical and sexual abuse of women and children we experience in South Africa. This is unacceptable for a society like ours where the rights of every citizen are entrenched in the constitution. Therefore, I have focused on the metaphor of the family in the Fourth Gospel, and how they are employed at micro-, meso- or macro-level. J.G. van der Watt’s book, Family of the King. Dynamics of Metaphor in the Gospel of John (2000) forms the basis of my study. He has pointed out that the metaphor of the family is the constitutive and most essential imagery in the Gospel. The story of the Samaritan woman in John 4 fits somehow into the family history of the father and the son. The questions I am concerned with are: 'How does Jesus' encounter with the Samaritan woman in John 4 fit into the network of imagery of the family in John's Gospel? What other imageries related to the imagery of the family, are also functioning in the story of the Samaritan woman and what significance does this encounter have in relation with the rest of the Gospel? In chapter 1 the research premise is worked out. Elements featuring in this chapter are: the problem statement, general and specific objectives, aim of the study, motivation and methodology. Literary criticism, social-scientific criticism, rhetorical criticism and theological criticism together are used into an integrated and approach to interpret this pericope. When they are used interactively, a rich and responsible approach is available for dealing with belief, action and life in the world today. In Chapter 2 the social-historical background of the Jewish and Roman family is discussed. Aspects like the meaning of family, family functions in the Jewish household and family and community solidarity were taking into consideration. The important role of cleanness and uncleanness in the Jewish family and the way they considered Samaritans as ‘menstuants from the cradle’ are underlined. Because John 4 is about the encounter between Jesus and a Samaritan woman, the origin of the Samaritans; their beliefs and traditions; and divorce in their community are important. The Samaritan education system; cleanness and uncleanness in their community and the place of a woman in the Samaritan family were also included in the discussion. At the end of this particular section the similarities and differences of the Jewish and Roman families were compared to be able to reconstruct and to obtain as much information as possible of the context of the Samaritan family. Chapter 3 contains the discussion of the meaning and function of metaphors in general and specific in the Gospel of John. In this study I have looked at the definition of a metaphor, types of metaphors and imagery in the Fourth Gospel. John emphasizes in his gospel the divinity of Jesus and his relationship with God. The author is using human relational images to portray this relationship as well as Jesus, the divine’s relationship to the world and to other people. In the words of Van den Heever: ‘The metaphors in John are all embedded in contexts made up by other metaphorical expressions: descent/ascent, living in you/you in me, partaking of Me as food, walking in the light, etcetera. It means that the connoted micro-level metaphors must be understood macro-metaphorically.’ (1992:94). This forms the basis of the discussion of the metaphor of the family in John 4. Chapter 4 is the focal point of the research and contains a detailed exegesis of John 4:1-42 in order to explain the functioning of the metaphor of the family on micro-level. The other metaphors that are linked with the metaphor of the family are also discussed, e.g. the metaphor of water and the significance of water in the Fourth Gospel as well as the metaphors of light and life. Because a family is about relationships, the family metaphors in John’s Gospel are about various relationships. It is in the first place about Jesus’ relationship with his Father, with the disciples and with the believers. In John 4 two other family relationships are portrayed: the Samaritan family and the Jewish family and they are in conflict with each other. Therefore, Jesus invites the Samaritan woman into a new family, namely the family of the Father. In this family she will have a special place and function. Other aspects of the Samaritan woman's relationships are also explored, e.g. her relationship with men, with the disciples and with men in the village. Jesus as a human being was also part of a family. The Gospel writers hesitated to say too much about it, but eventually we do know something about his family relationships. In the last chapter of my research families in South Africa are discussed. How the Fourth Gospel and particularly the story of the encounter between Jesus and the Samaritan woman can help to build families in South Africa that respects women; a society that gives women their rightful places in that society. / Dr. S.J. Nortje-Meyer

Magister Pharmaceuticae - MPharm / Plasmodium falciparum malaria continues to be a worldwide health problem, especially in developing countries in Africa and is responsible for over a million fatalities per annum. Chloroquine (CQ) is low-cost, safe and was the mainstay aminoquinoline derived chemotherapeutic agent that has been used for many years against blood-stage malaria. However, today the control of malaria has been complicated by increased resistance of the malaria parasite to existing antimalarial agents such as CQ. The primary cause of resistance is mutation in a putative ATP-powered multidrug efflux pump known as the p-glycoprotein (pGP) pump, and point mutation in P. falciparum CQ resistance transporter (PfCRT) protein. These mutations are responsible for the reduced accumulation of CQ at its primary site of action, the acidic digestive food vacuole of the parasite.To overcome the challenges of CQ resistance in P. falciparum, chemosensitiser offer an attractive approach. Chemosensitisers or reversal agents are structurally diverse molecules that are known to reverse CQ resistance by inhibiting the pGP efflux pump and/or the PfCRT protein associated with CQ export from the digestive vacuole in CQ resistant parasites. Chemosensitisers include the well-studied calcium channel blocker verapamil and antihistaminic agent chlorpheniramine. These drugs have little or no inherent antimalarial activity but have shown to reverse CQ resistance in P. falciparum when co-administered with CQ. Because of the channel blocking abilities of pentacycloundecylamines (PCUs) such as NGP1-01, it is postulated that these agents may act as chemosensitisers and circumvent the resistance of the Plasmodium parasite against CQ. Therefore as a proof of concept we conducted an experiment using CQ co- administered with different concentrations of NGP1-01 to evaluate the ability of NGP1-01 to act as a chemosensitiser.Herein, we report the ability of NGP1-01, the prototype pentacycloundecylamine (PCU), to reverse CQ resistance (> 50 %) and act as a chemosensitiser. NGP1-01 alone exhibited very low intrinsic antimalarial activity against both the resistant and sensitive strain (> 2000 nM), with no toxicity to the parasite detected at 10 µM. A statistically significant (p < 0.05) dose dependent shift was seen in the CQ IC50 values at both 1 µM and 10 µM concentration of co-administeredNGP1-01 against the resistant strain. Based on this finding we set out to synthesise a series of novel agents comprising of a PCU moiety as the reversal agent (RA) conjugated to a CQ-like aminoquinoline (AM) molecule and evaluate the potential of these PCU-AM derivatives as antimalarial- and/or reversed CQ agents. As recently shown by Peyton et al., (2012), the conjugation of a CQ-like molecule with a RA such as the chemosensitiser imipramine and derivatives thereof is a viable strategy to reverse CQ resistance in multidrug-resistant P. falciparum. The novel compounds were obtained by amination and reductive amination reactions. The synthetic procedures involved the conjugation of the Cookson’s diketone with different tethered 4-aminoquinoline moieties to yield the respective carbinolamines and the subsequent imines. This was followed by a transannular cyclisation using sodium cyanoborohydride as reducing agent to yield the desired PCU-AM derivatives. The CQ-like AMderivatives were obtained using a novel microwave (MW) irradiation method. Structure elucidation was done by utilising 1H- and 13C NMR spectroscopy as well as IR absorption spectrophotometry and mass spectrometry. Five PCU-AM reversed CQ derivatives were successfully synthesised and showed significant in vitro antimalarial activity against the CQ sensitive strain (NF54). PCU-AM derivatives 1.1 – 1.4 showed antimalarial IC50 values in the ranges of 3.74 – 17.6 ng/mL and 27.6 – 253.5 ng/mL against the CQ-sensitive (NF54) and CQ-resistant strains (Dd2) of Plasmodium falciparum, respectively. Compound 1.1 presented with the highest antimalarial activity against both strains and was found to be 5 fold more active against the resistant strain than CQ. The reversed CQ approach resulted in improved resistance reversal and a significantly lower concentration PCU was required compared to NGP1-01 and CQ in combination. This may be attributed to the improved ability of compound 1.1 to actively block the pGP pump and/or the increased permeability thereof because of the lipophilic aza-PCU moiety. Compound 1.1 also showed the lowest RMI value confirming that this compound has the best potential to act as a reversed CQ agent in the series. Cytotoxicity IC50 values observed for compounds 1.1 – 1.4 were in the low micromolar concentrations (2.39 – 9.54 µM) indicating selectivity towards P. falciparum (SI = 149 – 2549) and low toxicity compared to the cytotoxic agent emetine (IC50 = 0.061 µM).These results indicate that PCU channel blockers and PCU-AM derived conjugates can be utilised as lead molecules for further optimisation and development to enhance their therapeuticpotential as reversal agents and reversed CQ compounds.