This thesis describes the pathway of sulphur metabolism in Paracoccus denitrificans (NCIB 89MJ-) The compounds involved in the sulphur metabolic pathway were determined in growth experiments (Chapter 2) and time-course and pulse-chase experiments, using radioactively labelled sulphate (Chapter 3). P. denitrificans is an assimilatory sulphate-reducing organism with a pathway of sulphate reduction involving in- organic intermediates. P. denitrificans can not utilise cysteine as a sole sulphur; cysteine inhibits amino acid metabolism at low concentrations and respiration at higher concentrations. Sulphate is taken up by P. denitrificans against a concentration gradient (Chapter 4.) . The mechanism of sulphate uptake was investigated using right side out and inside out membrane vesicles prepared from P. denitrificans. The uptake mechanism involves an uncoupler-sensitive transport mechanism driven either by respiration, or by a transmembrane pH gradient (alkaline inside). The active transport of sulphate was shown to be carrier-mediated, by its sensitivity to sulphydryl-group reagents. It is proposed that the sulphate carrier operates by a mechanism of electroneutral proton symport, and is capable of transporting sulphate in either direction across the plasma membrane (Chapter 5). ATP Sulphurylase, the initial enzyme involved in the activation of sulphate, was purified, and its kinetic and regulatory properties investigated (Chapter 6). ATP Sulphurylase activity was repressed by sulphite, sulphide and cysteine, and inhibited by sulphide. Accumulation of APS, the end-product of the ATP sulphurylase-catalysed reaction, could only be detected in the presence of inorganic pyrophosphatase, an enzyme which removed pyrophosphate, another end-product of sulphate activation. Inorganic pyrophosphatase was purified and its substrate specificity, kinetics and regulatory properties examined, in relation to its part in sulphate metabolism (Chapter 7). Inorganic pyrophosphatase is a constitutive enzyme which functions equally well with either Mg²⁺ or Co²⁺ as the cofactor. APS Kinase activity was detected in crude extracts of P. denitrificans. A new assay is described for measuring APS kinase activity (Chapter 8). APS Kinase was purified. Coupled enzyme assays, with purified ATP sulphurylase, inorganic pyrophosphatase and APS kinase, indicated that all three enzymes were necessary for the synthesis, and accumulation of PAPS (Chapter 8). No 3andprime;-nucleotidase or enzyme "A" activity was detected. Serine transacetylase and O-acetyl serine sulphydrylase were purified and the kinetics and regulation of these two enzymes, investigated (Chapter 9). O-Acetyl serine lyase activity was detected in crude extracts of P. denitrificans, representing the first report of this enzyme in bacteria (Chapter 9). β-Cystathionase was purified and its kinetic and regulatory properties investigated; the unidirectionality of the cysteine to methionine pathway was confirmed (Chapter 10). Cysteinyl- and methionyl-tRNA synthetases were purified and the kinetics and regulation of these enzymes studied (Chapters 11 and 12 respectively). Both these enzymes possess different substrate specificities to the aminoacyl-tRNA synthetases from other organisms. Both enzymes appear to be constitutive. During this investigation of the sulphur metabolism of P. denitrificans, the substrate specificity of the different enzymes, to the selenium analogues of the respective sulphur-containing substrates, were investigated. Selenate competitively inhibits sulphate uptake and ATP sulphurylase, with respect to sulphate, but no APSe or PAPSe synthesis could be detected in the coupled enzyme assays (Chapters 7 and 8). Purified O-acetyl serine sulphydrylase catalysed the synthesis of selenocysteine from selenide and O-acetyl serine (Chapter 9). Both selenocysteine and selenomethionine are activated by the respective aminoacyl-tRNA synthetase (Chapters 11 and 12, respectively).

No description available.

The vascularly and luminally perfused rat jejunum has been developed as a useful experimental model for the study of intestinal function. However there are discrepancies in the reported results on the fate of luminally absorbed glucose in different versions of this system. In the present work, the vascularly and luminally perfused rat jejunum in vitro was established and thorough investigation was made of the dissection procedure and other experimental variables thought to be important in the functioning of the model. Special attention was paid to the preparation of the erythrocytes for the vascular perfusion medium to ensure their ability to pass through the vascular bed and to deliver oxygen to the intestinal tissues. Measurements of the respiration of the perfused intestine were made routinely in view of the paucity of such observations in the literature. Glucose absorption, translocation and metabolism by the intestine were measured using recirculation and once-through perfusion modes and, in the latter system, analysis of these functions was followed using both enzymatic and radiochemical assay techniques. Glucose and water absorption was also studied using a lumen-only recirculation perfusion of rat jejunum in vivo. The experiments were performed over a wide range of luminal glucose concentrations and osmolarities, and using proprietary glucose-electrolyte solutions intended for use in the oral rehydration of patients. In consequence of this work, it has been possible for the first time to realise why literature reports on the fate of luminally absorbed glucose differ so widely, and the present results appear to give the most accurate record so far on the distribution of this glucose (in the absence of other metabolisable substrates, and using this particular experimental system). It is concluded that the vascularly and luminally perfused rat jejunum in vitro appears to be the best available preparation for the study of the absorptive, translocation and metabolic functions of the small intestine.

The aim of these experiments was to investigate the effect of providing substrate in the form of CHO or fluid as water or a dilute glucose-electrolyte solution on the metabolic and circulatory changes during prolonged endurance exercise. It was hoped to establish the relative importance of fluid or substrate replacement in promoting endurance capacity. Subjects exercised significantly longer on a cycle ergometer when the glucose-electrolyte solution was given compared with the administration of large amounts of glucose or fructose in the form of polymer solutions. The effectiveness of the glucose-electrolyte solution in prolonging exercise time to exhaustion was not shown during exercise at high (33oC) and low (2oC) ambient temperatures. Fluid balance does not seem to be a priority during exercise at low ambient temperatures and exercise time in the heat was too short for the possible benefits of fluid replacement to occur. During 2h of moderate exercise (50% VO2max) at a high ambient temperature, the ingestion of the glucose-electrolyte solution was associated with the maintenance of plasma volume and minimal physiological disturbances which may limit performance. There was a tendency toward greater rectal temperatures and higher heart rates during rehydration with a hypertonic glucose polymer solution. The effect of fluid and substrate replacement during exercise performed over a range of intensities (50 - 70% VO2max) was investigated. It was suggested that the ingestion of a glucose polymer solution, in an attempt to provide glucose to the working muscle, may compromise fluid balance. The usefulness of carbohydrate feeding during prolonged exercise of moderate intensity where thermoregulation is a priority is questionable. The maintenance of plasma volume through the replacement of fluid losses during exercise seems to be the priority in the promotion of endurance capacity rather than the provision of carbohydrate.

Rate of oxygen consumption was used as a method of measuring the metabolic rate of flying locusts. The animals flew into an airstream from a wind tunnel while tethered to a balance so that lift could be determined. Wingbeat frequency was also recorded. The rate of oxygen consumption during flight was found to be determined principally by wingbeat frequency, but using mean values for steady flight good straight line correlations were also found between rate of oxygen consumption and lift, and wingbeat frequency and lift. However, the relationships between these three variables were seen to change during the early part of flight, when lift could be maintained while, in general, wingbeat frequency and rate of oxygen consumption declined. As wingbeat frequency decreased the amount of oxygen used with each wingbeat was found to remain fairly constant, but the amount of lift generated by each wingstroke increased. Therefore the proportion of the locust's total power output which was used to perform aerodynamic work was greater at the lower wingbeat frequency. It was considered that the action of adipokinetic hormone and the change from predominantly carbohydrate to lipid metabolism might be involved in the onset of the period of more economical, steady flight. However, no firm evidence of this was obtained from the experiments which were carried out. When locusts were injected with corpus cardiacum extract 1h before flight, their wingbeat frequency dropped more rapidly than that of saline injected animals. Injection of adipokinetic hormone immediately before flight had a detrimental effect on the locusts' flight performance.

No description available.

Call number: LD2668 .T4 1951 W45 / Master of Science

A dissertation submitted to the Faculty of Medicine University of the Witwatersrand, Johannesburg, in fulfilment of the requirements for the Degree Master of Science in Medicine / Ferritin is an iron storage protein, made up of heavy (H) and light (L) subunits. Ferritin synthesis is regulated at a post transcriptional level by iron, which induces a redistribution of ferritin mRNA from a free cytoplasmic pool to polyribosomes. Inflammatory states influence iron metabolism, causing a decrease in serum iron levels associated with an increase in reticuloendothelial ferritin synthesis and iron storage. / IT2018

A dissertation submitted to Faculty of Medicine in part fulfilment of the required for the degree of Master of Medicine in Psychiatry at the University of the Witwatersrand / Moclobemide is a reversible monoamine oxidase inhibitor. It is a short acting antidepressant, and previously studies were done comparing 300 to 600 mg dosages administered three times daily. Data on a twice daily dosage schedule is limited. This study compares the efficacy and tolerability of Moclobemide 150 mg twice daily as compared to two different three times a day schedules with total daily dosages of 300 to 450 mg respectively. Thirty one adult outpatients with major depressive disorder (as defined by DSM-IIIR) were included in this trial. It was controlled in a randomised double-blind manner. All treatments were found to be effective and no clinically significant differences between treatments could be demonstrated in this respect. Group 3 (150 mg b.d.) was found to be slightly more effective than the other two groups. Moclobemide was found to be effective at all dosage regimens with respect to treating co-morbid symptoms of anxiety and agitation. There were no differences between the groups with respect to type and frequency of adverse events. Overall tolerability was found to be good in all the treatment groups. Since the twice daily dosage is more convenient, it has been suggested that compliance will be enhanced on this regimen. The treatment response rates obtained confirm this conclusion. / IT2018

PhD - Science / Specialised mechanisms have evolved in pathogenic bacteria to enable adaptation to hostile and fluctuating host environments. Inducible mutagenesis, in particular, has been implicated in the emergence of antibiotic- and stress-resistant mutants. This thesis examined mycobacterial DNA metabolism with specific emphasis on the roles of multiple Y-family polymerases in the evolution of inter-strain variation and drug resistance in M. tuberculosis. The contribution of the nrdZ-encoded class II ribonucleotide reductase (RNR) to the maintenance of dNTP pools for replication and repair under hypoxic conditions was also explored. In addition, the co-factor requirement of NrdZ prompted an investigation into the biosynthesis and transport of adenosylcobalamin (AdoCbl) in M. tuberculosis. The data suggest that the mycobacterial Y-polymerases are tightly regulated and restricted to specialised damage-free repair or replication restart. Disruptions in individual M. smegmatis mc2155 DinB (pol IV) homologues resulted in novel antibiotic-resistance polymorphisms that were suggestive of non-redundant function. In contrast, abrogation of all error-prone polymerase activity failed to impair long-term competitive survival of mc2155 in vitro. Similarly, heterologous overexpression of M. tuberculosis pol IV homologues did not increase spontaneous mutation rates in wild-type mc2155, or complement damage hypersensitivity. However, treatment of M. smegmatis with gyrase inhibitors confirmed the differential induction of pol IV homologues in response to replication stalling and demonstrated elevated rates of spontaneous mutagenesis as a result of GyrB inhibition. The class II RNR does not appear to play a significant role in mycobacterial pathogenesis. Specifically, NrdZ was unable to substitute for the class I RNR under aerobic conditions in vitro, and a M. tuberculosis ÄnrdZ deletion mutant was not impaired in its ability to adapt to hypoxia in vitro. Similarly, infection of immunocompetent mice suggested that nrdZ is not required for the survival or virulence of M. tuberculosis in vivo. Disruptions in genes required for AdoCbl and methionine biosynthesis revealed that complex regulatory functions govern mycobacterial methionine and AdoCbl homeostasis. Loss of early (cobK) or late (cobU) stage AdoCbl biosynthetic enzymes had no effect on the growth of M. tuberculosis H37Rv in vitro. In contrast, deletion of the B12-independent methionine synthase (metE) resulted in impaired growth on solid media that could be rescued by vitamin B12 but not Lmethionine supplementation, simultaneously demonstrating the ability of M. tuberculosis to transport exogenous vitamin B12. Significantly, double ÄcobU/ÄmetE and ÄcobK/ÄmetE deletion mutants in which all predicted methionine synthase activity was eliminated, were not impaired for growth in liquid minimal media, suggesting that M. tuberculosis H37Rv possesses alternative mechanisms for methionine generation. Finally, the attenuated virulence of the ÄcobU and ÄmetE deletion mutants in vivo in immunocompetent mice indicated the relevance of AdoCbl biosynthesis to mycobacterial pathogenesis.