<p><font face="Times New Roman" size="3"><font face="Times New Roman" size="3"> <p align="left">The study made use of cross-sectional design that compared the performance of younger children (6- to 7-year-olds) with that of older children (12- to 13-year-olds) on various measures of executive function. Within this, it made use of a natural experimental design, with children exposed to alcohol<i><font face="Times New Roman" size="3"> as the experimental group and non-exposed children as the control group.</font></i></p> </font></font></p>

This thesis describes the target selection, cloning, expression, purification, crystallization, structure and biochemical characterization of five essential Mycobacterium tuberculosis (Mtb) proteins. The search for drugs against the causal agent of tuberculosis is urgently needed and the targeting of essential genes is necessary to fulfill this goal. The crystal structures of carbonic anhydrases (CA) Rv1284 and Rv3588c have been determined to 2.0 and 1.7 Å resolution, respectively. Rv3588c, in contrast to Rv1284, is an active β-CA that shows two different active site conformations and pH-dependent oligomerization states. Rv1295 is an active threonine synthase with an unusually high pH optimum; the structure has been solved to 2.5 Å resolution, based on which a modification to the reaction mechanism published previously is proposed. Mtb has a thick and impermeable cell envelope that constitutes an efficient barrier against drugs. One of the essential components of the envelope is mycolic acid (MA). The inhibition of enzymes participating in its synthesis would be lethal for Mtb. Rv0636, a formerly unknown-function protein has β-hydroxyacyl-ACP dehydrase activity which is essential for MA synthesis. Co-expression with partners notably improves its solubility. Around 55% of Mtb proteins have unknown function. Rv3778c is one of them and its three-dimensional structure has been determined to 1.8 Å resolution. Studies aimed at the elucidation of its biochemical function are shown. A pathway not yet reported in Mtb is also suggested.

The Fas pathway is critical for the maintenance of normal T cell homeostasis. Humans and mice with defects in this pathway exhibit the accumulation of large numbers of peripheral lymphocytes and lupus-like autoimmunity. A major feature of these organisms is the accumulation of non-NK TCRαβ+CD4-CD8- “double negative” (DN) T cells. While regulatory T cells (Tregs) with the DN phenotype have been extensively characterized in Fas-sufficient mice and humans, limited data exist on the role of DN T cells as Tregs in Fas-deficient animals. In fact, most of the literature suggests that the DN T cells accumulating in Fas-deficiency states are pathogenic, contributing to secondary lymph node enlargement and autoimmune disease. In this body of work, data are presented that illustrate that Fas-deficient lymphoproliferative (LPR) DN T cells can act as Tregs in an interferon γ (IFNγ)- and Fas ligand (FasL)-dependent fashion toward Fas-sufficient T cells. LPR DN T cells needed to be able to secrete and respond to IFNγ in order to upregulate surface FasL, in order to ameliorate GVHD mediated by CD4+ T cells in vivo and to suppress the proliferation of and kill activated CD4+ T cells in vitro. FcRγ, a key molecule involved in innate immune responses, can substitute for CD3ζ in the T cell receptor (TCR) of mouse and human T cells in certain circumstances; in doing so, it is essential for the regulatory function of TCR transgenic DN Tregs. FcRγ-deficient LPR mice were found to have exacerbated T cell accumulation and early mortality. We show that while FcRγ expression was required for LPR DN T cells to regulate CD4+ and CD8+ T cells responding to alloantigens in vitro and in vivo, it does not control autologous lymphoproliferation in LPR mice by supporting the function of a regulatory cell, nor does it affect the rate of proliferation of LPR T cells in vivo. Instead, FcRγ-expressing LPR CD4+, CD8+ and DN T cells were found to be undergoing apoptosis at a high rate in vivo, and in contrast to their FcRγ-deficient counterparts, FcRγ+ LPR DN T cells were capable of undergoing TCR restimulation-induced cell death (RICD). The data presented in this thesis therefore show that LPR DN T cells can exhibit IFNγ-, FasL- and FcRγ-dependent regulatory function, and also illustrate a previously unknown function for FcRγ in controlling the expansion of Fas-deficient T cells. The implications of these data for autoimmune lymphoproliferative syndromes, and normal T cell homeostasis, are discussed.

Previous research has shown a relationship between prenatal teratogenic exposure and impaired cognitive functioning. However, data regarding the long-term outcome of prenatal teratogenic exposure are minimal. The present study investigated the long-term neuropsychological functioning (specifically attention and memory) of adults prenatally exposed to alcohol or phenylalanine, and examined whether there was evidence to suggest that there are effects specific to individual teratogens. Using a battery of attention and memory measures the performance of 17 adults diagnosed with Fetal Alcohol Syndrome (FAS) and 13 adults with Maternal Phenylketonuria Syndrome (MPKUS) was assessed. In order to identify the pattern of deficits associated with prenatal teratogenic exposure, an age and CA and IQ matched control group was assessed. Attention was broadly assessed using Mirsky et al.'s (1991) neuropsychological model of attention. The memory and learning tests administered included a number of well standardized measures of verbal learning, verbal and visual recall, delayed recall, and recognition. Paired comparisons between the FAS group and age and CA and IQ matched controls indicated a unique pattern of attention and memory deficits consistent with previous research with children and adolescents. Specifically, adult individuals with FAS appear to have deficits in acquisition of new material, delayed recall of verbal material and in response inhibition. Paired comparisons between the MPKUS group and CA and IQ matched controls indicated that the pattern of attention and memory deficits seen in adults with MPKUS is difficult to distinguish when the effect of IQ is removed. A randomized block design using IQ as the blocking variable and group (FAS, MPKUS, or Controls) as the treatment variable was utilized to examine the question of whether the two prenatal teratogen groups differ from one another and from Controls in terms of attention and memory ability. Ten blocks of three participants (FAS, MPKUS and Control) matched on IQ were formed. The randomized block analyses revealed few differences between the groups and failed to reveal a number of the differences found in the paired comparisons between the prenatal teratogen groups and the CA and IQ matched Control group. Possible reasons for these differences are discussed.

This thesis entitled ‘ Unusual Acylation Properties of Type II Fatty Acid Biosynthesis Acyl Carrier Proteins’ describes the discovery of self-acylation and malonyl transferase activity in acyl carrier proteins involved in type II fatty acid biosynthesis and assigns a physiological role to these processes inside the cellular milieu. Acyl carrier protein (ACP) is one of the most abundant proteins present inside the cell and almost 4% enzymes require it as a cofactor. Acyl carrier proteins can exist either as discrete proteins or as domains of large functional proteins. They function in a variety of synthases as central molecules to which growing acyl intermediates and nascent product molecules are covalently tethered during the elongation and modification steps required to produce the final product. A prototypical bacterial ACP is composed of 70-80 amino acids and is generally expressed in the apo form. It is post-translationally modified to active holo form by the addition of 4'-phosphopantetheine moiety to an absolutely conserved serine residue in a reaction catalyzed by holo-ACP synthase or 4'-phosphopantetheine transferase. Chapter 1 surveys literature related to carrier proteins inside the cell and describes the thesis objective. It also presents an overview of the acyl carrier proteins and their involvement in various metabolic pathways inside the cell. The chapter details the structural organization of acyl carrier proteins from various sources revealing the conservation in their structure and also details the molecular basis of interaction of ACP with other enzymes inside the cell. The discovery of unusual self-acylation property in acyl carrier proteins involved in polyketide biosynthesis and its absence in acyl carrier proteins involved in fatty acid biosynthesis prompted me to investigate the reasons for this selective behavior. Discovery of self-acylation property in acyl carrier proteins Plasmodium falciparum and chloroplast targeted Brassica napus acyl carrier proteins involved in type II fatty acid biosynthesis and the mechanism of this reaction forms the basis of Chapter 2. In this chapter it has been shown that self-acylation property is intrinsic to a given acyl carrier protein and is not dependent on the pathway in which it is involved. Based on primary sequence analysis and site directed mutagenesis studies presence of an aspartate/glutamate has been identified to be critical for the self-acylation event. Furthermore, it has also been shown that the self-acylation event in type II fatty acid biosynthesis acyl carrier proteins is highly specific in nature employing only dicarboxylic acid –CoAs as substrates unlike the polyketide biosynthesis acyl carrier proteins which utilize both dicarboxylic acid and β-keto acid thiol ester -CoAs as substrates. The detailed kinetics of these reactions has also been worked out. Combining all the results a plausible mechanism for the self-acylation reaction has been proposed. Chapter 3 describes the discovery of a novel malonyl transferase behavior in acyl carrier proteins involved in type II fatty acid biosynthesis. Malonyl transferase property in ACPs of type II FAS from a bacterium (Escherichia coli), a plant (Brassica napus) and a parasitic protozoon (Plasmodium falciparum) were investigated to present a unifying paradigm for the mechanism of malonyl transferase behavior in ACPs. Identification of malonyl transferase property in Plasmodium falciparum ACP and Escherichia coli ACP (EcACP) and the absence of this property in Brassica napus ACP has been described in this chapter. Detailed investigations demonstrated that presence of an arginine or a lysine in loop II and an arginine or glutamine at the start of helix III as the residues that are critical for the transferase activity. In order to assign a physiologic function to these unusual acylation properties, fabD(Ts) mutant strain of Escherichia coli was utilized for heterologous complementation by the various wild type and mutant ACPs that are able to catalyze either or both of the activities. Growth of the mutant strain at non-permissive temperature, when complemented with ACPs catalyzing both the reactions confirmed that these properties have a physiologic relevance. Extensive mutagenesis experiments in conjunction with complementation studies allowed me to propose a plausible mechanism on how the self-malonylation and malonyl transferase properties operate in tandem. Chapter 4 describes the thermodynamic characterization of self-acylation process using Isothermal Titration Calorimetry. Isothermal Titration Calorimetric studies on the binding of malonyl, succinyl, butyryl and methylmalonyl –CoA to Plasmodium falciparum and Brassica napus acyl carrier proteins were performed to investigate the role of thermodynamic parameters in the specificity of self-acylation reaction. Calculation of the parameters showed that the thermodynamics does not control the self-acylation reaction. The evolution of self-acylation property in various acyl carrier proteins and its possible significance in the evolution of various metabolic events is described in Chapter 5. Extensive bioinformatics search was performed and phylogenetic analysis on acyl carrier proteins from 60 different taxa was done using the MEGA4 program. Analysis showed that this property was first found in cyanobacterium. Later, during the course of evolution this property was lost in most acyl carrier proteins, and was retained either in acyl carrier proteins that are targeted to organelles of cyanobaterial orgin viz. apicoplast in apicomplexans and chlorplasts in plants or in acyl carrier proteins involved in secondary metabolic events such as polyketide biosynthesis. Chapter 6 summarizes the findings of the thesis. Acyl carrier protein from Plasmodium falciparum, Brassica napus and Escherichia coli were characterized for their self-acylation and malonyl transferase properties and a combined mechanism for these two properties is proposed. The work done also provides an in vivo rationale to these in vitro processes. Furthermore, the evolutionary significance of the self-acylation behavior is also discussed in the thesis. The thesis also probes into the thermodynamics of the self-acylation reaction in Plasmodium falciparum and Brassica napus acyl carrier proteins. Thus, the thesis adds a new dimension to the much unexplored ACP biology and paves the way to study in vivo roles of these processes in detail. Appendix I describes the Isothermal Titration calorimetric characterization of binding of various acyl-PO4 molecules to Escherichia coli PlsX (Acyl-phosphate acyltransferase). PlsX, the first enzyme of phosphatidic acid biosynthesis pathway catalyzes the conversion of acyl-ACP into acyl-PO4, which is further used by other enzymes leading to the formation of phosphatidic acid. ITC results presented in this section show that longer chain length acyl-PO4 molecules show better binding to PlsX, as compared to the smaller ones demonstrating that long chain acyl molecules serve as better substrates for phosphatidic acid synthesis.

Renal Cell Carcinoma (RCC) has low 5 year survival rate and is resistant to radiation and chemotherapy. Phenethyl Isothiocyanate (PEITC) is a naturally occurring phytochemical that has a variety of anti-cancer properties. Here we explore two anti-cancer properties of PEITC: induction of apoptosis and induction of cell cycle arrest in RCC cells and the underlying mechanisms. We used two human RCC cell lines Caki-1 and Caki-2. Survival and cell proliferation was assayed using Calcein AM. Annexin V staining was used to measure apoptosis. Caspase-3/7 induction was measured using a fluorescent substrate. Cell cycle was studied using Propidium Iodide staining. DNA damage was determined using phospho [gamma]-H2AX antibody. Protein expression and phosphorylation was determined using immunoblotting. PEITC significantly reduced survival of Caki-1 and Caki-2 cells and inhibited their proliferation as determined by Calcein AM. 15 and 20 [mu]M PEITC induced apoptosis in both cell lines and induced caspase-3/7 activity. Western blot analysis revealed caspase-8, caspase-9 and Bid cleavage as well as upregulation of the death receptors Fas and DR5. Lower doses (up to 10 [mu]M) arrested Caki-1 cells in G2/M phase, and this was associated with increased p38 and MK2 (Thr334) phosphorylation. The p38 inhibitor SB203850 inhibited this G2 arrest induced by PEITC. 15 and 20 [mu]M PEITC treatment resulted in increased [gamma]-H2AX phosphorylation suggesting DNA damage, but this was completely blocked by caspase inhibitor. In summary, our study shows that PEITC induces apoptosis in Caki-1 and Caki-2 cells by upregulating Fas and DR5 and activating the downstream apoptosis cascade. PEITC does not cause direct DNA damage to the cells; the observed DNA damage is a result of the apoptotic process and is blocked by caspase inhibitor. PEITC induces G2/M arrest in Caki-1 cells and the mechanism involves p38 phosphorylation which activates MK2. Inducing cell cycle arrest and apoptosis may play an important role in the anti-cancer properties of PEITC. Fully understanding the mechanism by which PEITC induces apoptosis and cell cycle arrest in RCC cells may lead to development of novel chemotherapeutic drugs against RCC. / text

The purpose of this report is to evaluate the hardness and microstructure in bent and welded samples of Alloy 718. The results will be used by GKN aerospace to evaluate the simulated values of the production process of vines in a jet engine. In total, eleven samples from three different production chains are evaluated. All samples are bent and go through different stages within the production including bending, solution treatment and welding. The samples are cut and mounted in bakelite then polished and etched in order for the microstructure to be seen and evaluated. Hardness tests were made on the mounts to evaluate how bending, solution treatment and welding affects the hardness. The bent samples without the solution treatment became harder depending on the amount of cold deformation. The amount of cold deformation controls how fast the material recrystallizes during subsequent solution treatment. During the solution treatment, the δ-phase (Ni3Nb) is precipitated in the grain boundaries which prevents a coarser grain size and therefore promotes a smaller grain size.  The laser weld creates a small HAZ (heat affected zone) that becomes softer because the heat dissolves the δ-phase which therefore triggers the grains to grow. The weld consists of eutectic γ-dendrites with interdendritic pools of alloying elements. After the solution treatment, many needle shaped δ-phases arise from the pools and HAZ. The hardness measurements were tested on a manual machine which makes potential human error important to consider when the measurements are evaluated. When the grain size measurements are done on the solution treated samples, the grain boundaries can be difficult to determine because the δ-phases and twins create wide and incorrect boundaries. The results will be used by GKN Aerospace in order to verify their simulations. The heterogenic material after solution treatment will probably be studied further. / Syftet med projektet är att undersöka hårdhet och mikrostruktur i bockade och svetsade prover tillverkade av Alloy 718. Totalt undersöks elva stycken prover som är tillverkade på olika sätt enligt tre tillverkningskedjor. Alla provbitar är bockade och har sedan genomgått olika många steg i tillverkningen som består utav bockning, upplösningsbehandling och svets. Genom att kapa upp provbitarna i mindre bitar, baka in de i bakelit och etsa de studerades mikrostrukturen och hur den påverkas av bockning, upplösningsbehandling och svets. Vidare gjordes hårdhetsmätningar över olika delar av proven för att undersöka hur mycket hårdare materialet blir vid kalldeformation och hur upplösningsbehandling och svets påverkar hårdheten i Alloy 718. Endast bockade prov blir hårdare beroende på hur mycket kalldeformation provbiten har utsatts för. Hur mycket provet har deformerats styr hur snabbt rekristallisationen sker vid en efterföljande upplösningsbehandling. Under upplösningsbehandlingen utskiljs även δ-fas (Ni3Nb) i korngränser vilket främjar en mindre kornstorlek. Lasersvetsen ger ett litet värmepåverkat område men värmen från svetsen bidrar till att δ-fasen löses upp och större korn bildas. Svetsen består av långa eutektiska γ-dendriter med interdendritiska poler av mycket legeringsämnen. Vid upplösningsbehandling efter utförd svets bildas det δ-fas i de värmepåverkade och i svetsen. Hårdhetsmätningarna har utförts på en manuell hårdhetsmaskin vilket innebär att felmarginalen blir större då den mänskliga felfaktorn spelar en stor roll. Vid beräkning av kornstorlek har det varit svårt att utskilja vad som är en korngräns i upplösningsbehandlade prov då mängden δ-fas efter upplösningsbehandling ger tjocka korngränser.  Resultaten kommer användas av GKN Aerospace för att verifiera sina simulationer. Det heterogena materialet efter upplösningsbehandlingen kommer troligtvis studeras vidare.

Mental retardation is a heterogeneous group of disorders with countless causes. It is characterised by cognitive and functional limitations in everyday skills, for example social skills, communication skills and motor skills and can be classified in behavioural, etiological and educational systems. Down's syndrome and Fetal Alcohol Syndrome are two of the many syndromes defined under mental retardation. The goal of this dissertation was to determine the effect of a water activity intervention programme on the motor proficiency levels of children with Down's syndrome and Fetal Alcohol Syndrome. These aims were addressed by structuring the dissertation in five chapters: Chapter one constituting the introduction and statement of the problem, Chapter 2 presenting a review of relevant literature, Chapters 3 and 4 consisting of two research articles, addressing the specific aims of the study, and Chapter 5 including the summary, conclusions and recommendations. All the children who participated in the study were intuitionalized in a school for the mentally and physically handicapped. The MABC-test was used as the main evaluation instrument, and components of the Charlop-Atwell test were used to evaluate the coordination skills of the children with Down's syndrome. The first aim of this study was to determine the effect of a specially designed water activity intervention programme on the motor proficiency levels of children with Down's syndrome. Six children classified as having Down's syndrome, formed part of the research group. Their chronological age ranged between 9 and 14 years while their mental age classification was that of a 4 to 5 year old. The data was analysed by means Summary of descriptive statistics, and effect sizes were determined. The second aim of the study was to determine the effect of a water activity intervention programme on the motor proficiency levels of children with Fetal Alcohol Syndrome. Six children participated in the programme. Their chronological age ranged between 7 and 17 years while their mental age classification was that of a 4 to 11 year old. Reporting the results were in the form of case studies, and effect sizes of differences were determined. With regard to the first aim of the study the results indicated that the motor proficiency levels of the experimental group with Down's syndrome improved, especially regarding the MABC-total, balance- and total body coordination skills. With reference to the second aim of the study, the results indicated that improvement in the motor proficiency levels of the children with Fetal Alcohol Syndrome had a lasting effect. The MABC total, ball skills and manual dexterity were the components that showed the best improvement. It can be concluded that a water activity intervention programme is a suitable method for rectifying motor deficiencies among children with Down's syndrome and Fetal Alcohol Syndrome. Recommendations for the improvement of the water activity programme were presented, as well as suggestions for further studies. / Thesis (M.A. (Human Movement Science))--North-West University, Potchefstroom Campus, 2004.

<p><font face="Times New Roman" size="3"><font face="Times New Roman" size="3"> <p align="left">The study made use of cross-sectional design that compared the performance of younger children (6- to 7-year-olds) with that of older children (12- to 13-year-olds) on various measures of executive function. Within this, it made use of a natural experimental design, with children exposed to alcohol<i><font face="Times New Roman" size="3"> as the experimental group and non-exposed children as the control group.</font></i></p> </font></font></p>

Mental retardation is a heterogeneous group of disorders with countless causes. It is characterised by cognitive and functional limitations in everyday skills, for example social skills, communication skills and motor skills and can be classified in behavioural, etiological and educational systems. Down's syndrome and Fetal Alcohol Syndrome are two of the many syndromes defined under mental retardation. The goal of this dissertation was to determine the effect of a water activity intervention programme on the motor proficiency levels of children with Down's syndrome and Fetal Alcohol Syndrome. These aims were addressed by structuring the dissertation in five chapters: Chapter one constituting the introduction and statement of the problem, Chapter 2 presenting a review of relevant literature, Chapters 3 and 4 consisting of two research articles, addressing the specific aims of the study, and Chapter 5 including the summary, conclusions and recommendations. All the children who participated in the study were intuitionalized in a school for the mentally and physically handicapped. The MABC-test was used as the main evaluation instrument, and components of the Charlop-Atwell test were used to evaluate the coordination skills of the children with Down's syndrome. The first aim of this study was to determine the effect of a specially designed water activity intervention programme on the motor proficiency levels of children with Down's syndrome. Six children classified as having Down's syndrome, formed part of the research group. Their chronological age ranged between 9 and 14 years while their mental age classification was that of a 4 to 5 year old. The data was analysed by means Summary of descriptive statistics, and effect sizes were determined. The second aim of the study was to determine the effect of a water activity intervention programme on the motor proficiency levels of children with Fetal Alcohol Syndrome. Six children participated in the programme. Their chronological age ranged between 7 and 17 years while their mental age classification was that of a 4 to 11 year old. Reporting the results were in the form of case studies, and effect sizes of differences were determined. With regard to the first aim of the study the results indicated that the motor proficiency levels of the experimental group with Down's syndrome improved, especially regarding the MABC-total, balance- and total body coordination skills. With reference to the second aim of the study, the results indicated that improvement in the motor proficiency levels of the children with Fetal Alcohol Syndrome had a lasting effect. The MABC total, ball skills and manual dexterity were the components that showed the best improvement. It can be concluded that a water activity intervention programme is a suitable method for rectifying motor deficiencies among children with Down's syndrome and Fetal Alcohol Syndrome. Recommendations for the improvement of the water activity programme were presented, as well as suggestions for further studies. / Thesis (M.A. (Human Movement Science))--North-West University, Potchefstroom Campus, 2004.