The 33 kDa manganese stabilizing protein (MSP) has been proposed to provide ligands to stabilize Mn ions in the water lysis reaction of photosystem II of photosynthesis. In previous research site-directed mutagenesis had been performed on regions of the psbO gene encoding two aspartic acid residues of MSP which were thought to have the potential to form carboxyl bridges with Mn ions. The purpose of this research was to analyze these mutants. Plasmids pUC120-33 (#1,3,5,7,9,11,15) containing mutant psbO genes could not be isolated from E.coli because the expressed MSP was toxic to the cells. However, a psbO mutant gene carried in pPGV5-33 (#7) was isolated from E.coli and transformed into cyanobacterium Svnechococcus PCC 7942. Cyanobacterial cells carrying the MSP mutant showed a susceptibility to intensive light (100 footcandles) with a decrease of 30% in the growth rate within the first 100 hours after inoculation. This result suggested a possible function of the MSP in protecting the oxygen evolving complex from intensive light exposure. However, the mutant appeared to revert after this time probably due to homologous gene recombination with the wild type gene. In order to further analyze the function of mutants without recombination occurring, the construction of an MSP deletion was attempted using insertion of a kanamycin cartridge into the middle of the psbO gene. The inactivated psbO gene was transformed into E.coli and transformants were selected by kanamycin resistance. However, plasmid DNA carrying the interrupted genes could not be isolated, probably due to toxicity of the expression product in E.coli cells. Thus, future studies should be directed to reconstruction of a deletion mutant by direct transformation into cyanobacterial cells. Once a deletion mutant has been constructed analyses of the site-directed mutations could be performed in cyanobacteria. / Department of Biology

Electron spin echo envelope modulation (ESEEM) spectroscopy is widely used to study the radical pairs created during the primary steps of photosynthesis. In this thesis the analysis of ESEEM spectra is improved, and some new applications and variations of this experiment suggested. Experimental spectra from species such as P⁺Q^-_A, the secondary radical pair formed in the reaction centre of the bacterium Rhodobacter sphaeroides, give information about the exchange and dipolar couplings between the radicals. The model used to analyse the data affects the results; this thesis suggests two improvements. First, the effect of anisotropic hyperfine couplings in the radicals is considered by the addition of a single spin-1/2 nucleus to the model. This approach suggests that previous models neglecting the effect of nuclei may have been slightly in error. Secondly, several model fittings are performed in the time domain. This approach avoids the Fourier transformation to the frequency domain so that experimental dead-time does not corrupt the data. An excellent fit to experimental data is found with a model containing one spin-1/2 nucleus on each radical. The hyperfine coupling parameters resulting from the fit are consistent with independent experimental results. Use is made of the method of Cramér-Rao lower bounds to assess the precision to which experimental parameters are determined from a time domain curve fitting. It is shown that the lower bounds may also be used to determine the optimum sampling strategy for the experiment. An example is given of the novel use of ESEEM to determine the distance between the radicals in the strongly coupled, uncorrelated radical pair Q^-_AQ^-_B ESEEM has not yet been used for this purpose, and the simulated spectra produced here indicate that the experiment could be used to evaluate the dipolar coupling and hence the inter-radical distance. This thesis considers the possibility of performing ESEEM at higher frequencies than are usually considered. Calculations show that the increased resolution of the g-tensors allow an experiment performed at the W-band frequency of 95 GHz to make a correlation between the relative orientations of the radicals and the dipolar axis, information which has previously been unavailable from a single experiment.

Purple nonsulfur bacteria are a group of extraordinary metabolic diverse bacteria. They can grow photoautotrophically, photoheterotrophically , chemoheterotrophically or chemoautotrophically. Under various conditions, they can enjoy exceptional flexibility within each of these modes of metabolism. Due to the special physical characteristics properties, they had attracted scientist¡¦s attention in resent years. These bacteria are widely distributed in nature such as lakes, water ponds, coastal lagoons or high concentration organic waste lagoons. Lotus Pond, located in northern Kaohsiung City, is a serious eutrophied artificial lake. Because of receiving sufficient light and having been polluted by significant amounts of soluble organic matter, the ecology of the lake is suitable for the growth of purple nonsulfur bacteria. In the study, the lake water and sediments by using a Winograsdsky column, we successfully isolated 16 strains bacteria from the Lotus Pond. We also amplified the 16S-rDNA fragments of these strains by PCR and sequenced these PCR products, then aligned these sequences with the data of GeneBank. We affirmed that the 16 isolated strains belong to purple nonsulfur bacteria. From phylogenetic analysis, these 16 strains belong to the following three groups of bacteria: Rhodopseudomonas palustris, Rubrivivax gelatinosus, and Rhodobacter sphaeroides. Characteristic studies of these strains, we found that all isolated strains are Gram negative bacteria and contain bacteriochlorophyll a. The strains that belong to R. palustris and R. sphaeroides group can use several different types of short chain organic acid as their carbon source and have denitrification ability. However, only the strains belong to R. palustris group are able to use the aromatic compound benzoate. From salt tolerant studies, we found the strains in R. sphaeroides group can grow well in 3% NaCl, and both R. palustris and R. gelatinosus group can only grow in 1% NaCl.

Photobiological hydrogen production from organic materials occurs with the help of illumination and under aerobic conditions within photobioreactors. Novel designs are needed in order to increase the light conversion efficiency and to improve the biological hydrogen production. In this thesis, purple non sulfur bacteria Rhodobacter sphaeroides O.U. 001 was employed as the hydrogen producing microorganism. Two different types of photobioreactors, namely oscillatory helical photobioreactor and recycling helical bioreactor, were devised and successfully operated for bacterial growth and hydrogen production. Total liquid capacity of the pneumatically driven oscillatory flow helical tubular photobioreactor was 11.5 L, and 4.5 L of which was occupied by the bacterial culture. The bacteria grew very well both in malate-based and acetate-based media under nitrogen atmosphere. The bacteria sustained their vitality 24 days before the system was shut down. The recycling helical tubular photobioreactor, which was developed for hydrogen production, had a fully occupied total volume of 6.5L. The bacteria produced approximately 1.9L of hydrogen in four days on malate-based media. The hydrogen production rate was 0.009LH2/Lculture.h. The effects of molecular nitrogen gas and the sodium glutamate concentration on the growth of hydrogen producing photosynthetic bacteria Rhodobacter sphaeroides O.U.001 in the reactor were also examined in 500ml-bottles. The bacterial growth curves did not show any difference at the control medium containing 15mM of acetate and 10 mM of sodium glutamate. However, other bottles containing a lesser amount of N-source was found to grow earlier under the nitrogen atmosphere. Besides, even a 15/2 acetate/sodium glutamate ratio was observed to be sufficient to grow the bacteria for inoculation, and to spend extra sodium glutamate was not necessary. The novel designs developed in this study aim to improve the biological hydrogen production by photosynthetic bacteria, and to provide new ways in adaptation of photobiological systems to outdoor conditions for large-scale applications.

Heavy metals are one of the most important environmental pollutants. In recent years, many low cost stretages of bioremediation for contaminated sites by heavy metals, such as fungi, bacteria and plants have been investigated for their biosorption capacity towards heavy metals. The uses of plant species for remediate contaminated sites by heavy metals are so called phytoremediation. The purpose of the first parts of this study are to (1) evaluate bioavailability of Cadmium (Cd) in contaminated soil and phytoremediation potential by three plant species, Vetiveria zizanioides, Pteris multifida, and Alternanthera philoxeroides (Mart.), and (2) realized the influence of photosynthetic bacteria (PSB) on the uptake of Cd in the three species. The results showed that the Alternanthera philoxeroides (Mart.) could accumulate the highest concentration of Cd among the three species, in which the Cd concentration of plant tissue increased with the concentration in soil. The highest concentration of Cd (164.9 mg kg-1) was found in the below-ground parts of Alternanthera philoxeroides (Mart.) at the 8th week of culturing period. However, the species of Vetiveria zizanioides could accumulate the largest total Cd, up to 547.5 £gg/ plant, which thus extracted the greatest amounts of Cd from the soil. Therefore, in the first part of this study the species of Vetiveria zizanioides was concluded to be the best accumulator among the three plant species. In addition, the concentration of Cd in the species of Pteris multifida was found significantly increased after PSB was added into the soil, but the plants died later due to Cd stress. The experimental results also showed that PSB seemed to be not suitable for each species used in this study to accumulate Cd from Cd-contaminated soil. In the second part of this research, both pot and field experiments were conducted to (1) evaluate bioavailability of copper (Cu), lead (Pb) and zinc (Zn) in contaminated soil and phytoremediation potential by domesticated plants, Bidens pilosa and Passiflora foetida inoculated with arbuscular mycorrhizal (AM) fungi, and to (2) compare the results of pot and field experiments. The plant species of Bidens pilosa inoculated with AM fungi had significantly higher Cu concentrations in the shoots and roots than non-inoculated plants. The plant species of Passiflora foetida inoculated with AM fungi also had significantly higher Cu and Pb concentrations in the roots than non-inoculated plants. When we found that the root dry weight of Passiflora foetida inoculated with AM fungi dramatically increased, the concentrations of Cu, Pb and Zn in the root of the plant species increased by 9-14 times, comparing with the plants without inoculation of AM fungi. The AM fungi have potential either to promot plant growth or to increase heavy metal accumulation. The values of element translocation proportion from root to shoot was Zn>Cu>Pb for the plant species in both pot and field experiments. For both experiments, the results of pot test and field test were significantly different. The concentrations of pot tests were found higher than the field tests, and some values of pot tests were even found significantly greater than those in the field tests. In the third part of this study, the field experiments were conducted to test the feasibility of using domesticated vegetations for phytoremediation of the contaminated farmland. The objectives of this study were (1) to acquire information about the ability of five plant species growing wild in the polluted area to accumulate Cu, Pb and Zn, (2) to investigate the season effects on phytoremediaton of five plant species and evaluate the total uptake of heavy metal, and (3) to run both pot tests and a field trial of phytotremediation to compare their differences. The experimental results showed that three maximum toxic elements in a pot were 3020 mg kg-1 Pb, 232 mg kg-1 Cu and 1012 mg kg-1 Zn respectively. The Cu concentrations of the five plant species collected from the polluted plots ranged from 0.7 to 17.43 mg kg-1. The range of variation of Pb in plant tissues was measured varied from 2.29 to 81.65 mg kg−1, while a wide range of Zn concentrations was found from 12.84 to 192.85 mg kg-1 among the plants collected at the contaminated plots. In comparison to winter season, the Zn concentrations in Broussonetia papyrifera, Passiflora foetida and Saccharum sinensis collected in summer season was significantly higher. The higher Cu concentrations were obtained in both plant species of Bidens pilosa and Mimosa diplotricha in summer season. However, Pb concentrations in Saccharum sinensis collected in winter were significantly higher than those in the same plant species collected in summer. Bidens pilosa was also found having the highest total amount of Cu and Zn. The highest total amount of Pb was found in Mimosa diplotricha. For both plant species, both of the pot and field tests were different.

Thesis (Ph. D.)--University of California, San Diego, 2000. / Vita. Includes bibliographical references.

Photosynthetic membranes were prepared from the cyanobacterium Synechococcus PCC 7942 with oxygen evolving specific activity of 250-300 µmoles 02/ mg chl/hr. The membranes retained activity with a half-life of 4-5 days when stored at 0°C, or when quickly frozen in liquid nitrogen, greater than 95% of the activity remained after 2 months. Attempts to purify homogeneous preparations of photosystem II complexes from these membranes by detergent extraction were unsuccessful as indicated by a lack of a significant increase in oxygen evolution specific activity of the detergent extracts. Photosynthetic membrane detergent extracts usually maintained the same oxygen evolution specific activity as the orginal membranes, and a considerable amount of Photosystem I activity (75 µmoles 02 consumed /mg chl/hr in the Mehler reaction) was still present. The donor side of the photosystem II particles in the detergent extract was intact since the artificial electron acceptor, 2,6-dichiorophenolindophenol (DCPIP), was reduced at a rate comparable to the oxygen evolving activity. All oxygen evolving activity of the detergent extracts was lost when ion-exchange chromatography was used to resolve the co-extracted photosystem II and photosystem I complexes. / Department of Biology

Typescript (Photocopy) / xxx, 296 leaves : ill. ; 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 Agricultural Biochemistry, 1983

Thesis (Ph.D.) -- University of Adelaide, Dept. of Agricultural Biochemistry, 1983. / Typescript (Photocopy).

Thesis (M.Phil.) - University of Queensland, 2006. / Includes bibliography.