The production of engineered nanoparticles (ENPs) is growing at an incredibly fast rate and will soon become a trillion dollar industry. At this rate of production, there is a great potential for engineered nanomaterials to be released into the environment, both intentionally and unintentionally. TiO2 ENPs are one of the most widely produced nanoparticles with a broad range of applications in paints, inks, sunscreens, cosmetics, astronautics, and air/water purification. TiO2 ENPs have been proposed for their use in agricultural settings as a UV protectant, a defense against harmful bacteria and fungi, or a catalyst for the degradation of pesticides and herbicides. Furthermore, it has been shown to increase several aspects of photosynthesis in spinach including Rubisco and Rubisco activase activity, chlorophyll synthesis, and oxygen evolution. Foliar application of TiO2 ENPs on spinach resulted in a significant increase in plant fresh weight, dry weight, chlorophyll content, net photosynthetic rate, and carboxylase activity of Rubisco. These findings have prompted investigations for the use of TiO2 ENPs as a foliar spray to promote plant growth and yield. The first main objective of this research was to determine if TiO2 ENPs has the capabilities to increase photosynthetic production in Zea mays at concentrations similar to that of the experiments performed with spinach. Secondly, it was examined if the size of the TiO2 was a factor in the increased photosynthetic response by comparing TiO2 ENPs with bulk TiO2. Finally, the determination of whether the boost in photosynthesis resulted in an increased seed quality/quantity. Another aspect of this research was to determine how the interaction of TiO2 ENPs with inorganic contaminants may affect the uptake and accumulation of the contaminants in plants. Cadmium and arsenic are two of the top ten most hazardous substances on the priority list of the Agency for Toxic Substances and Disease Registry. Sources for Cd and As contamination include atmospheric deposition resulting from mining, smelting, and fuel combustion, phosphate fertilizers, and sewage sludge. Both of these contaminants can be taken up by plant roots and translocated to the leaves and fruits, thus entering the food chain. The release of TiO2 ENPs into domestic and industrial wastewaters is expected to represent the largest release of these nanoparticles. There has been data showing that up to 99% of TiO2 ENPs that enter wastewater treatment plants are retained in the sludge. In addition, TiO2 ENPs are being used at some water treatment plants because of their strong adsorption strength for hazardous materials, such as cadmium, arsenic, and copper and also the photocatalytic breakdown of harmful organic compounds. Since sewage sludge from wastewater treatment plants is applied to agricultural lands as a soil conditioner and fertilizer, this has resulted in the introduction of an estimated 120 g kg-3 per year of TiO2 ENPs. With sewer sludge being the common factor for contamination of agricultural fields, there is a high potential for the simultaneous introduction of TiO2 ENPs and heavy metal contaminants. To date, there has been very little research done for ENP and contaminant interactions. Of the research that has been performed on the subject, the majority of it was conducted using aquatic systems involving fish and daphnids. This research has shown that the interaction of TiO2 ENPs and metal contaminants generally increases the concentration of the contaminant in the organism, however it is still unclear whether the contaminant is biologically available or if it is adsorbed to the surface of the TiO2 ENPs. This information gives rise to two alternative hypotheses on how TiO2 ENPs may affect the fate of heavy metal contaminants in a single substrate growth media. The first is that the TiO2 ENPs may sequester the heavy metals in the soil thus decreasing the amount of the heavy metals that can be taken up by the plant. The alternative is that the TiO2 ENPs could act as a carrier of the metals i.e. if the plant is able to take up the intact TiO2 ENP with heavy metals adsorbed to the surface, it could potentially increase the amount of the metals that enter the plants. The main objective of this study was to determine which of these scenarios is true for broccoli plants that were grown in cadmium and arsenate contaminated growth media.

Tomatoes (Lycopersicon esculentum L.) require high amounts of nitrogen to maximize fruit production. The type of nitrogen and timing of fertilizer applications are important in tomato production systems to reduce nitrogen losses while optimizing yields. A two-year greenhouse study was conducted to determine the effects of organic and inorganic fertilization treatments on nitrogen and carbon isotopic compositions of tomato plant leaves and fruit at immature and mature stages of plant growth. The carbon and nitrogen isotopic ratios together will also help to better depict trends that develop from conventional versus organically grown tomatoes and their resulting water use efficiency (WUE). ‘Better Bush’ tomatoes were grown in 8 L plastic pots filled with 1:1:1 steamed-sterilized sand:silt loam soil:peat mix. Eight fertility treatments were evaluated: no fertility, synthetic Miracle Grow® (MG), organic bonemeal and bloodmeal (BB), BB with liquid Earthjuice (BBL), BB with 25% vermicompost (VC), BBL with 25% VC, MG with 25% VC, and no fertility with 25% VC. The results indicated that for both growth stages, δ15NAir differed (P ≤ 0.05) between fertility treatments, while no major differences were observed for δ13CVPDB (P > 0.05), although conventionally fertilized tomatoes with MG and MG with 25% VC tended to have lower δ13CVPDB values suggesting a greater transpirational water loss through open stomata. The organic treatments with VC had higher δ15NAir values than the conventional or no added fertility treatments for all five leaflets from three different branches, fruit skins from the first two fruit clusters, fruit juices from the first two clusters and soil samples. Generally, the 15N/14N stable isotope values of tomato foliage and fruit are distinctly different between organic and conventional fertilizers, which could provide a powerful forensic tool in fingerprinting tomatoes grown by organic farming methods. Therefore, nitrogen isotopes can distinguish among different fertility treatments and also help to label the VC applications, which are thought to provide better fertility management due to less soil leaching and volatilization.

abstract: The Heliobacterial reaction center (HbRC) is generally regarded as the most primitive photosynthetic reaction center (RC) known. Even if the HbRC is structurally and functionally simple compared to higher plants, the mechanisms of energy transduction preceding, inside the core, and from the RC are not totally established. Elucidating these structures and mechanisms are paramount to determining where the HbRC is in the grand scheme of RC evolution. In this work, the function and properties of the solubilized cyt c553, PetJ, were investigated, as well as the role HbRC localized menaquinone plays in light-induced electron transfer, and the interaction of the Nif-specific ferredoxin FdxB with reaction center particles devoid of bound FA/FB proteins. In chapter 2, I successfully express and purify a soluble version of PetJ that functions as a temperature dependent electron donor to P800+. Recombinant PetJ retains the spectroscopic characteristics of membrane-bound PetJ. The kinetics were characteristic of a bimolecular reaction with a second order rate of 1.53 x 104 M-1s-1 at room temperature and a calculated activation energy of 91 kJ/mol. In chapter 4, I use reverse phase high-performance liquid chromatography (HPLC) to detect the light-induced generation of Menaquinol-9 (MQH2) in isolated heliobacterial membranes. This process is dependent on laser power, pH, temperature, and can be modified by the presence of the artificial electron acceptor benzyl viologen (BV) and the inhibitors azoxystrobin and terbutryn. The addition of the bc complex inhibitor azoxystrobin decreases the ratio of MQ to MQH2. This indicates competition between the HbRC and the bc complex, and hints toward a truncated cyclic electron flow pathway. In chapter 5, the Nif-Specific ferredoxin FdxB was recombinantly expressed and shown to oxidize the terminal cofactor in the HbRC, FX-, in a concentration-dependent manner. This work indicates the HbRC may be able to reduce a wide variety of electron acceptors that may be involved in specific metabolic processes. / Dissertation/Thesis / Doctoral Dissertation Biochemistry 2015

abstract: To mimic the membrane environment for the photosynthetic reaction center of the photoheterotrophic Heliobacterium modesticaldum, a proteoliposome system was developed using the lipids found in native membranes, as well as a lipid possessing a Ni(II)-NTA head group. The liposomes were also saturated with menaquinone-9 to provide further native conditions, given that menaquinone is active within the heliobacterial reaction center in some way. Purified heliobacterial reaction center was reconstituted into the liposomes and a recombinant cytochrome c553 was decorated onto the liposome surface. The native lipid-attachment sequence of cytochrome c553 was truncated and replaced with a hexahistidine tag. Thus, the membrane-anchoring observed in vivo was simulated through the histidine tag of the recombinant cytochrome binding to the Ni(II)-NTA lipid's head group. The kinetics of electron transfer in this system was measured and compared to native membranes using transient absorption spectroscopy. The preferential-orientation of reconstituted heliobacterial reaction center was also measured by monitoring the proteoliposome system's ability to reduce a soluble acceptor, flavodoxin, in both whole and detergent-solubilized proteoliposome conditions. These data demonstrate that this proteoliposome system is reliable, biomimetic, and efficient for selectively testing the function of the photosynthetic reaction center of Heliobacterium modesticaldum and its interactions with both donors and acceptors. The recombinant cytochrome c553 performs similarly to native cytochrome c553 in heliobacterial membranes. These data also support the hypothesis that the orientation of the reconstituted reaction center is inherently selective for its bacteriochlorophyll special pair directed to the outer-leaflet of the liposome. / Dissertation/Thesis / Masters Thesis Chemistry 2018

abstract: The heliobacterial reaction center (HbRC) is widely considered the simplest and most primitive photosynthetic reaction center (RC) still in existence. Despite the simplicity of the HbRC, many aspects of the electron transfer mechanism remain unknown or under debate. Improving our understanding of the structure and function of the HbRC is important in determining its role in the evolution of photosynthetic RCs. In this work, the function and properties of the iron-sulfur cluster FX and quinones of the HbRC were investigated, as these are the characteristic terminal electron acceptors used by Type-I and Type-II RCs, respectively. In Chapter 3, I develop a system to directly detect quinone double reduction activity using reverse-phase high pressure liquid chromatography (RP-HPLC), showing that Photosystem I (PSI) can reduce PQ to PQH2. In Chapter 4, I use RP-HPLC to characterize the HbRC, showing a surprisingly small antenna size and confirming the presence of menaquinone (MQ) in the isolated HbRC. The terminal electron acceptor FX was characterized spectroscopically and electrochemically in Chapter 5. I used three new systems to reduce FX in the HbRC, using EPR to confirm a S=3/2 ground-state for the reduced cluster. The midpoint potential of FX determined through thin film voltammetry was -372 mV, showing the cluster is much less reducing than previously expected. In Chapter 7, I show light-driven reduction of menaquinone in heliobacterial membrane samples using only mild chemical reductants. Finally, I discuss the evolutionary implications of these findings in Chapter 7. / Dissertation/Thesis / M.S. Biochemistry 2012

M.Ed. (Didactics - Natural Sciences) / The specific aim of this study is to identify misconceptions about· photosynthesis among Standard 9 pupils. To identify the possible misconceptions which may occur about photosynthesis, the writer made an empirical study using questionnaires which comprised of definitions, multiple choice and descriptive questions. The possible misconceptions and the possible nature of the misconceptions about photosynthesis were identified after the completion of the questionnaire. Comparisons were then made to see whether there was a correlation between the relevant study and other studies in the literature available.

Global modeling of gross primary production (GPP) is a critical component of climate change research. On local scales, GPP can be assessed from measuring CO₂ exchange above the plant canopy using tower-based eddy covariance (EC) systems. The limited footprint inherent to this method however, restricts observations to relatively few discrete areas making continuous predictions of global CO₂ fluxes difficult. Recently, the advent of high resolution optical remote sensing devices has offered new possibilities to address some of the scaling issues related to GPP using remote sensing. One key component for inferring GPP spectrally is the efficiency (ε) with which plants can use absorbed photosynthetically active radiation to produce biomass. While recent years have seen progress in measuring ε using the photochemical reflectance index (PRI), little is known about the temporal and spatial requirements for up-scaling these findings continuously throughout the landscape. Satellite observations of canopy reflectance are subject to view and illumination effects induced by the bi-directional reflectance distribution function(BRDF) which can confound the desired PRI signal. Further uncertainties include dependencies of PRI on canopy structure, understorey, species composition and leaf pigment concentration. The objective of this research was to investigate the effects of these factors on PRI to facilitate the modeling of GPP in a continuous fashion. Canopy spectra were sampled over a one-year period using an automated tower-based, multi-angular spectroradiometer platform (AMSPEC), designed to sample high spectral resolution data. The wide range of illumination and viewing geometries seen by the instrument permitted comprehensive modeling of the BRDF. Isolation of physiologically induced changes in PRI yielded a high correlation (r²=0.82, p<0.05) to EC-measured ε, thereby demonstrating the capability of PRI to model ε throughout the year. The results were extrapolated to the landscape scale using airborne laser-scanning (light detection and ranging, LiDAR) and high correlations were found between remotely-sensed and EC-measured GPP (r²>0.79, p<0.05). Permanently established tower-based canopy reflectance measurements are helpful for ongoing research aimed at up-scaling ε to landscape and global scales and facilitate a better understanding of physiological cycles of vegetation and serve as a calibration tool for broader band satellite observations. / Forestry, Faculty of / Graduate

This thesis reports results of a study of relationships between photosynthetic activity and developmental parameters, and cambial activity (wood formation rate), during and following the period of active wood formation in untreated white spruce seedlings, and in seedlings stressed either by extensive stem incisions, or by tilting. The approach involved the use of two non-destructive methods for measuring photosynthetic activity: chlorophyll a fluorescence using optical instrumentation, and CO₂ uptake using infrared gas exchange techniques. Photosynthetic development was examined by estimating chlorophyll a content from a specific fluorescence parameter (O-level), and by the relative occurence of specific chloroplast stroma and membrane (thylakoid) proteins using electrophoretic and immunoblotting techniques. Cambial activity was determined using digitized image analysis of prepared cross-sections of seedling stems. Several fluorescence parameters were strongly correlated to cambial activity in untreated seedlings during the period of active wood formation (in mid-summer). However, the correlations were severely diminished or non-existent when cambial activity was arrested (in late-summer and autumn). Correlations between fluorescence and cambial activity in stressed seedlings were not discernible at any time, suggesting that the induced stresses resulted in a substantial alteration in normal source:sink relationships. Carbon dioxide uptake measures, either uncorrected or corrected to estimated chlorophyll α content, were not measurably correlated to cambial activity in untreated or stressed seedlings at any time in this system. Chlorophyll α content estimated from O-level fluoresecence, was not related to cambial activity in untreated or stressed seedlings. The relative occurences of two enzymes and proteins associated with photosynthetic carbon fixation, i.e. ribulose 1,5-bisphosphate carboxylase (Rubisco) and Coupling Factor, did not appear to be influenced by applied wounding and geotropic stresses. In contrast to the strong correlations found between fluorescence parameters and current season stem vigour, pre-season seedling height and cross-sectional stem areas were not related to stem vigour. These results suggest that in unstressed white spruce seedlings, the measure of specific chlorophyll α fluorescence parameters, using the methods delineated in this study, offers an alternative and more strongly predictive means of assessing current stem vigour, than measures of seedling dimensions. The results of this study provide strong evidence for, and a degree of elucidation on, the anticipated but previously unestablished existence of a source:sink relationship between leaves and vascular cambium in conifer seedlings. This information should provide an initial foundation for the elucidation of non-invasive methodologies by which to assess stem vigour of white spruce seedlings, and to probe source:sink relationships in other conifer species. / Science, Faculty of / Botany, Department of / Graduate

Rhodobacter capsulatus photosynthesis gene was isolated by creating in-frame fusions in a lacZ transcriptional/translational vector, and selecting for those that directed oxygen-regulated levels of β-galactosidase in R. capsulatus. One lacZ fusion isolate was used to identify an open reading frame (ORF) of unknown function and flanking sequences that promoted initiation of transcription. Interposon mutagenesis experiments identified the ORF as the bchC gene, which encodes an enzyme that catalyses the penultimate step in the biosynthesis of bacteriochlorophyll a, and also showed that the bchC gene formed an operon with the bchA gene. The nucleotide sequence of this bchC gene and its 5' regulatory region were determined. The deduced amino acid sequence showed that the bchC gene encodes a 33 kDA protein that has hydrophobic segments that could interact with a lipid membrane, and that this putative BchC protein contains a potential bacteriochlorophyll a binding site. Deletion analysis, S1-nuclease protection, and primer extension experiments showed that promoter activity was associated with sequences to which a 5' end mapped, and that these sequences had significant similarity to the proposed promoter regions of several other R. capsulatus photosynthesis genes. RNA blotting and S1-nuclease protection end-mapping experiments using bipartite probes provided direct evidence that the mRNA transcripts of the bchCA operon overlap those of the two flanking operons, the crtEF and the puf operons, such that the crtEF, bchCA, and puf operons may be cotranscribable, and that RNA polymerase may initiate transcription at one of several promoters. The significance of these overlapping mRNAs was evaluated using two interposon mutant strains, one that prevented crtEF transcripts from overlapping those the bchCA and puf operons, and the other that prevented both crtEF and bchCA transcripts from overlapping those of the puf operon. The results suggested that transcriptional readthrough stimulates promoter activity. Moreover, a pufB::lac'Z fusion could be expressed from the bchCA promoter equally as well as from the puf promoter, suggesting that these overlapping transcripts are functionally significant in the chromosomal context. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

The primary amino acid sequences of the L, M, and H photosynthetic reaction center peptide subunits from a number of purple non-sulfur bacteria, including Rhodopseudomonas viridis, Rhodobacter sphaeroides, and Rhodobacter capsulatus have been previously shown to be highly homologous, and detailed X-ray crystallographic analyses of reaction centers from two species of purple non-sulfur bacteria, Rps. viridis and R. sphaeroides have shown that all recognized structural and functional features are conserved. Experiments were undertaken to determine whether genes encoding reaction center and light harvesting peptide subunits from one species could be functionally expressed in other species. Plasmid-borne copies of R sphaeroides and Rps. viridis pigment binding-peptide genes were independently introduced into a photosynthetically incompetent R. capsulatus mutant host strain, deficient in all known pigment-binding peptide genes. The R. sphaeroides puf operon, which encodes the L and M subunits of the reaction center as well as both peptide subunits of light harvesting complex I, was shown to be capable of complementing the mutant R. capsulatus host. Hybrid reaction centers, comprised of R. sphaeroides-encoded L and M subunits and an R. capsulatus-encoded H subunit, were formed in addition to the R. sphaeroides-encoded LHI complexes. These hybrid cells were capable of photosynthetic growth, but their slower growth rates under low light conditions and their higher fluorescence emission levels relative to cells containing native complexes, indicated an impairment in energy transduction. The Rps. viridis puf operon was found to be incapable of functional expression in the R. capsulatus mutant host. Introduction of a plasmid-borne copy of the Rps. viridis puhA gene, which encodes the H subunit of the reaction center, into host cells already containing the Rps. viridis puf operon, such that all structural peptides of the Rps. viridis reaction center were present, still did not permit stable assembly of Rps. viridis photosynthetic complexes. RNA blot analysis demonstrated that the barrier to functional expression was not at the level of transcription. Differences between Rps. viridis and R. sphaeroides that may account for their differing abilities to complement the R. capsulatus mutant host strain are discussed. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate