No description available.

No description available.

Among the various fungal strains screened for citric acid production, Aspergillus niger is known to produce considerable amounts of citric acid and other organic acids when cultivated in carbohydrate-rich medium in solid substrate fermentation (SSF). Since A. niger on a solid substrate grows under conditions similar to the natural habitat, SSF is ideal to cultivate A. niger for the purpose of producing citric acid. / An initial optimization (study 1) was conducted in batch type fermentation experiments using peat moss supplemented with glucose to simulate an organic waste. The effects of various nutrients (glucose, (NH4)2SO 4, KH2PO4 and NaCl), fermentation parameters (moisture content, temperature, inoculum density, composition of solid substrate and particle size) and of initial level of potential stimulators (ethanol, methanol, phytate and surfactant) were evaluated with respect to citric acid production by A. niger grown on damp peat moss. In these experiments, optimization using a traditional 'one-factor-at-a-time' method was applied to determine key factor ranges for the production of citric acid. When the fermentation was carried out using the final optimal conditions. This allowed for a 50-fold increase in citric acid production compared to the production of citric acid by A. niger grown on peat moss supplemented with 100 g glucose/kg DPM. / A second set of experiments (study 2) was conducted to optimize fermentation conditions for citric acid production in a column bioreactor. A. niger NRRL 567 grown on damp peat moss was held within a column bioreactor and periodically irrigated with a glucose-rich solution simulating field conditions. Three variables including aeration, thickness of solid substrate bed and incubation temperature were optimized using a 23 full factorial design (FFD). Under optimum, the total citric acid production and yield were 120.6 g/kg DPM and 18.5% respectively. / A third experiment (study 3) compared the production of citric acid by A. niger in submerged fermentation using cheese whey, as opposed to batch and semi-continuous fermentation using peat moss. Various fermentation conditions such as nutrients (glucose, (NH4)2SO 4 and KH2PO4), stimulators (methanol, olive oil and phytate) and fermentation parameters (pH, fermentation time and inoculum density) were optimized using a central composite design (CCD). Citric acid production improved citric acid production by a factor of 13.3 when compared to the production of citric acid by A. niger NRRL 567 using whey-based medium (50 g/l) alone. / As compared to submerged and semi-continuous fermentation, Batch type SSF could take higher levels of initial glucose and produce the high concentration of citric acid within a shorter period of time. Thus, SSF may be considered to be better technique than submerged fermentation, if main disadvantage like non-homogeneous fermentation conditions could be overcome.

xi, 165 leaves : photos., graphs ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Pathology, 1979

vii, 136 leaves : ill., graphs, tables, photos ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.1979) from the Dept. of Biochemistry, University of Adelaide

Abstract In eukaryotic cells, fatty acids are mainly synthesized in the cytoplasm, but recently, in yeast and in humans, the ability to synthesize fatty acids has been characterized in mitochondria. This mitochondrial pathway is similar to bacterial type II fatty acid synthesis (FAS). The main feature of mitochondrial FAS in yeast is the respiratory deficient phenotype and loss of cytochromes when any of genes encoding enzymes for mitochondrial FAS is deleted. Mitochondrial FAS has been demonstrated to have an important role in lipoic acid production, namely it synthesizes octanoyl-ACP, which is used as a precursor for lipoic acid. However, the role and function of mitochondrial FAS is not yet fully understood. Many components of the mitochondrial FAS pathway in yeast have been identified according to their similarity to bacterial counterparts, but 3-hydroxyacyl-ACP dehydratase does not show any easily recognizable similarity to bacterial dehydratases and thus remained unidentified. In this study 3-hydroxyacyl-ACP dehydratases of mitochondrial FAS were characterized from the yeast Saccharomyces cerevisiae, humans, and the human pathogen Trypanosoma brucei. The yeast 3-hydroxyacyl-ACP dehydratase (Htd2p) was identified by using a genetic screen, and this protein was shown to be encoded by open reading frame (ORF) YHR067w. The product of this gene shows mitochondrial localization and exhibits hydratase 2 activity. The deletion of HTD2 leads to a respiratory deficient phenotype, loss of cytochromes, reduced lipoic acids levels and changes in mitochondrial morphology. The ORF encoding human 3-hydroxyacyl-ACP hydratase (HsHTD2) was identified by functional complementation of the respiratory deficient phenotype of the yeast htd2 mutant with a human cDNA library. The complementing cDNA was previously identified as the RPP14 transcript encoding the 14 kDa subunit of the human RNase P complex. It was found that this transcript contains another 3' ORF, which encodes a protein that displays hydratase 2 activity and has mitochondrial localization. The bicistronic nature of the transcript is conserved in vertebrates and indicates a genetic link between mitochondrial FAS and RNA processing. The mitochondrial 3-hydroxyacyl-ACP hydratase in T. brucei is homologous to human HTD2, can complement the yeast respiratory deficient phenotype, exhibits hydratase 2 activity and localizes to the T. brucei mitochondrion.

Abstract Fatty acid degradation in living organisms occurs mainly via the β-oxidation pathway. When this work was started, it was known that the hydration and dehydrogenation reactions in mammalian peroxisomal β-oxidation were catalyzed by only multifunctional enzyme type 1 (MFE-1; Δ2-Δ3-enoyl-CoA isomerase/2-enoyl-CoA hydratase 1/(3S)-hydroxyacyl-CoA dehydrogenase) via the S-specific pathway, whereas in the yeast peroxisomes via the R-specific pathway by multifunctional enzyme type 2 (MFE-2; 2-enoyl-CoA hydratase 2/(3R)-hydroxyacyl-CoA dehydrogenase). The work started with the molecular cloning of the rat 2-enoy-CoA hydratase 2 (hydratase 2). The isolated cDNA (2205 bp) encodes a polypeptide with a predicted molecular mass of 79.3 kDa, which contains a potential peroxisomal targeting signal (AKL) in the carboxyl terminus. The hydratase 2 is an integral part of the cloned polypeptide, which is assigned to be a novel mammalian peroxisomal MFE-2. The physiological role of the mammalian hydratase 2 was investigated with the recombinant hydratase 2 domain derived from rat MFE-2. The protein hydrates a physiological intermediate (24E)-3α, 7α, 12α-trihydroxy-5β-cholest-24-enoyl-CoA to (24R, 25R)-3α, 7α, 12α, 24-tetrahydroxy-5β-cholestanoyl-CoA in bile acid synthesis. The sequence alignment of human MFE-2 with MFE-2(s) of different species reveals 12 conserved protic amino acid residues, which are potential candidates for catalysis of the hydratase 2. Each of these residues was replaced by alanine. Complementation of Saccharomyces cerevisiae fox-2 (devoid of endogenous MFE-2) with human MFE-2 provided a model system for examing the in vivo function of the variants. Two protic residues, Glu366 and Asp510, of the hydratase 2 domain of human MFE-2 have been identified and are proposed to act as a base and an acid in catalysis. Mammalian MFE-2 has a (3R)-hydroxyacyl-CoA dehydrogenase domain, whereas the yeast MFE-2 has two dehydrogenase domains, A and B. The present work, applying site-directed mutagenesis to dissect the two domains, shows that the growth rates of fox-2 cells expressing a single functional domain are lower than those of cells expressing S. cerevisiae MFE-2. Kinetic experiments with the purified proteins demonstrate that domain A is more active than domain B in catalysis of medium- and long-chain (3R)-hydroxyacyl-CoA, whereas domain B is solely responsible for metabolism of short-chain substrates. Both domains are required when yeast cells utilize fatty acids as the carbon source.

No description available.

No description available.

No description available.