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A genetic suppressor approach to the biogenesis, quality control and function of photosynthetic complexes in Chlamydomonas reinhardtiiMalnoë, Alizée 08 July 2011 (has links) (PDF)
Central in oxygenic photosynthesis, the cytochrome b6f complex, couples electron transfer to proton translocation across the thylakoid membrane via its quinol:plastocyanin oxidoreductase activity, contributing to ATP formation. Cytochrome b6f complex differs from its respiratory homolog, the bc1 complex, by the presence of an additional heme, heme ci located within the quinone reduction site Qi and attached by a unique thioether bond. Mutants lacking heme ci show low accumulation of partially functional b6f complex and, hence, cannot grow phototrophically. This grounded a screen for suppressor mutations that would restore higher accumulation of b6f complexes whose function, even if compromised, would sustain phototrophic growth.The genetic suppressor approach undertook in Chlamydomonas reinhardtii during this PhD thesis led to the isolation and characterisation of the ftsh1-1 protease mutant (mutation R420C which should affect ATP hydrolysis). The mutant ftsh1-1 proved to be a versatile tool for the functional study of otherwise degraded proteins. The combination of genetic, biochemical, physiological and biophysical experiments demonstrated notably that: (i) a QiKO mutant, whose b6f complexes are devoid of both bh and ci hemes, can grow phototrophically despite a broken Q-cycle, (ii) the absence of covalently bound heme ci, in the Rccb2 mutant, triggers photosensivity enhanced in the presence of O2 supporting a role for heme ci in oxygen rich environment, (iii) FtsH is involved in the maintenance of the main photosynthetic complexes.
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Substrate water binding to the oxygen-evolving complex in photosystem IINilsson, Håkan January 2014 (has links)
Oxygenic photosynthesis in plants, algae and cyanobacteria converts sunlight into chemical energy. In this process electrons are transferred from water molecules to CO2 leading to the assembly of carbohydrates, the building blocks of life. A cluster of four manganese ions and one calcium ion, linked together by five oxygen bridges, constitutes the catalyst for water oxidation in photosystem II (Mn4CaO5 cluster). This cluster stores up to four oxidizing equivalents (S0,..,S4 states), which are then used in a concerted reaction to convert two substrate water molecules into molecular oxygen. The reaction mechanism of this four-electron four-proton reaction is not settled yet and several hypotheses have been put forward. The work presented in this thesis aims at clarifying several aspects of the water oxidation reaction by analyzing the mode of substrate water binding to the Mn4CaO5 cluster. Time-resolved membrane-inlet mass spectrometric detection of flash-induced O2 production after fast H218O labelling was employed to study the exchange rates between substrate waters bound to the Mn4CaO5 cluster and the surrounding bulk water. By employing this approach to dimeric photosystem II core complexes of the red alga Cyanidoschyzon merolae it was demonstrated that both substrate water molecules are already bound in the S2 state of the Mn4CaO5 cluster. This was confirmed with samples from the thermophilic cyanobacterium Thermosynechococcus elongatus. Addition of the water analogue ammonia, that is shown to bind to the Mn4CaO5 cluster by replacing the crystallographic water W1, did not significantly affect the exchange rates of the two substrate waters. Thus, these experiments exclude that W1 is a substrate water molecule. The mechanism of O-O bond formation was studied by characterizing the substrate exchange in the S3YZ● state. For this the half-life time of this transient state into S0 was extended from 1.1 ms to 45 ms by replacing the native cofactors Ca2+ and Cl- by Sr2+ and I-. The data show that both substrate waters exchange significantly slower in the S3YZ● state than in the S3 state. A detailed discussion of this finding lead to the conclusions that (i) the calcium ion in the Mn4CaO5 cluster is not a substrate binding site and (ii) O-O bond formation occurs via the direct coupling between two Mn-bound water-derived oxygens, which were assigned to be the terminal water/hydroxy ligand W2 and the central oxo-bridging O5. The driving force for the O2 producing S4→S0 transition was studied by comparing the effects of N2 and O2 pressures of about 20 bar on the flash-induced O2 production of photosystem II samples containing either the native cofactors Ca2+ and Cl- or the surrogates Sr2+ and Br-. While for the Ca/Cl-PSII samples no product inhibition was observed, a kinetic limitation of O2 production was found for the Sr/Br-PSII samples under O2 pressure. This was tentatively assigned to a significant slowdown of the O2 release in the Sr/Br-PSII samples. In addition, the equilibrium between the S0 state and the early intermediates of the S4 state family was studied under 18O2 atmosphere in photosystem II centers devoid of tyrosine YD. Water-exchange in the transiently formed early S4 states would have led to 16,18O2 release, but none was observed during a three day incubation time. Both experiments thus indicate that the S4→S0 transition has a large driving force. Thus, photosynthesis is not limited by the O2 partial pressure in the atmosphere.
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Limitation of photosynthetic carbon metabolism in South African soybean genotypes in response to low night temperatures / Abram Johannes StraussStrauss, Abram Johannes January 2008 (has links)
Thesis (Ph.D. (Botany))--North-West University, Potchefstroom Campus, 2009.
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The hydrogen-bonded water network in the oxygen-evolving complex of photosystem IIPolander, Brandon C. 13 January 2014 (has links)
Protein dynamics play a key role in enzyme-catalyzed reactions. Vibrational spectroscopy provides a method to follow these structural changes and thereby describe the reaction coordinate as a function of space and time. A vibrational spectroscopic technique, reaction-induced FTIR spectroscopy, has been applied to the study of the oxygen-evolving complex (OEC) of photosystem II (PSII). In plant photosynthesis, PSII evolves oxygen from the substrate, water, by the accumulation of photo-oxidizing equivalents at the OEC. Molecular oxygen and protons are the products of this reaction, which is responsible for the maintenance of an aerobic atmosphere on earth. The OEC is a Mn4CaO5 cluster with nearby bound chloride ions. Sequentially oxidized states of the OEC are termed the S states. The dark-stable state is S1, and oxygen is released on the transition from S3 to S0. Using short laser flashes, individual S states are generated, allowing vibrational spectroscopy to be used to study these different oxidation states of the OEC. In current X-ray crystal structures, hydrogen bonds to water molecules are predicted to form an extensive network around the Mn4CaO5 cluster. In the OEC, four peptide carbonyl groups are linked to the water network, which extends to two Mn-bound and two Ca-bound water molecules. This dissertation discusses a vibrational spectroscopic method that uses these peptide carbonyl frequencies as reporters of solvatochromic changes in the OEC. This technique provides a new, high-resolution method with which to study water and protein dynamics in PSII and other enzymes.
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Electrical characterization of microwire-polymer assemblies for solar water splitting applicationsYahyaie, Iman 03 1900 (has links)
The increasing demand for energy and the pressure to reduce reliance on fossil fuels encourages the development of devices to harness clean and renewable energy. Solar energy is a large enough source to fulfill these demands, however, in order to overcome its daily and seasonal variability, it has been proposed that sunlight be harvested and stored in the form of chemical fuels. One potential approach is the photosynthetic splitting of water to store solar energy in the simplest chemical bond, H–H, using a device that includes: semiconducting microwire arrays as light harvesting components, redox catalysts, and a membrane barrier for separating the products of water redox reactions..
However, the harvested solar energy can be lost across the system and it is critical to characterize the electrical properties of each component within the system to quantify how much of this energy will ultimately be coupled to the water splitting reactions. The aim of this research is to develop approaches for characterization of a proposed system of this kind, incorporating individual semiconductor microwires as photoelectrodes (with no redox catalysts) embedded into a candidate conducting polymer membrane to form a single functional unit.
Semiconductor microwires were isolated and using a novel contact formation approach with tungsten probes in a standard probe station, and their current versus voltage properties were characterized. This approach is of particular interest when
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considering the limitations of conventional contact formation approaches (e.g. thermal evaporation of contact metals), arising from the small dimensions of the microwires and also the incompatibility of these techniques with many microwire/polymer structures due to the unwanted interactions between polymers, photoresists, etchants and the high temperature lithographic processes.
The electrical properties of different microwires and also the junctions between microwires and two candidate polymers were studied. Specifically, the combination of methyl-terminated silicon microwires and PEDOT:PSS:Nafion demonstrated promising behavior, with a total DC resistance of approximately 720 kΩ (i.e. losses < 16 mV at maximum available photocurrent), making it a suitable candidate for the use in the proposed system. The outcome of these research may be applied to many applications including semiconducting microstructures and conducting polymers.
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Limitation of photosynthetic carbon metabolism in South African soybean genotypes in response to low night temperatures / Abram Johannes StraussStrauss, Abram Johannes January 2008 (has links)
Thesis (Ph.D. (Botany))--North-West University, Potchefstroom Campus, 2009.
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Internal leaf CO₂ transfer conductance diffusional limitation and its consequences for modelling photosynthesis in C₃ plant speciesEthier, Gilbert J. 10 March 2010 (has links)
Virtually all current estimates of the maximum carboxylation rate (V.) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and the maximum electron transport rate (.Imax) for C3 species used to parameterise Land Surface Models (LSMs) implicitly assume an infinite CO2 transfer conductance from intercellular spaces to the sites of carboxylation (gi). And yet, most measurements in perennial plant species or in ageing or stressed leaves show that gi imposes a significant limitation on photosynthesis. In this study, I demonstrate that many current parameterisations of the photosynthesis model of Farquhar, von Caemmerer & Berry (1980) based on the leaf intercellular CO, concentration (Ci) are incorrect for leaves where gi limits photosynthesis. I show how conventional A-C, curve (net CO2 assimilation rate of a leaf - An - as a function of Ci) fitting methods which rely on a rectangular hyperbola model under the assumption of infinite gi can significantly underestimate Vcmax for such
leaves. Alternative V., parameterisations of the conventional method based on a single, apparent Michaelis-Menten constant for CO, evaluated at C; and used for all C3 plants are also found to be inaccurate since the relationship between Vcmax and g; is not conserved among species. To address this problem, I present an alternative curve fitting method that accounts for gi through a non-rectangular hyperbola version of the model of Farquhar et al. (1980).
Current estimates of a central Farqhuar et al. (1980) model parameter, Kc(1+O/K0) (effective Michaelis-Menten constant for CO2), vary 4-fold, making it very difficult to justify any single value for the parameterisation of large scale, pan-species LSM studies. Following on previous work published over two decades ago, I demonstrate that the current range of Kc(1+O/K0) values chosen for LSMs is partly an artefact of many inaccurate in vitro determinations, and results in widely different estimates of An for given Vcmac values. Once corrected, the average Kc(1 +O/Ko) value determined in vitro for C, plants is essentially identical to the two in vivo values published to date, but considerable variation within the data set remains due to the poor accuracy of the in vitro determinations. The new A-Ci curve fitting method elaborated in this study suggests new ways of obtaining in vivo estimates of Rubisco's kinetic constants, as I demonstrate through a well-documented example.
The CO, transfer conductance was originally considered to be a constitutional property of a leaf related to its internal anatomy. This study provides the first estimates of gi in a coniferous species and examines variation in gi through time and space in relation to anatomical and physiological traits. Gas exchange measurements and subsequent novel A-Ci curve analyses, as well as stable carbon isotope, nitrogen (N), protein, and pigment analyses, were made on upper and lower canopy, current- to 4-year-old needles of 50-year-old Pseudotsuga menziesii trees. During the first growing season, needle thickness and leaf mass per area decreased by 30% from the top to bottom of the canopy. These anatomical changes were accompanied by modest variation in area-based estimates of g , but no causal link could be established between anatomical traits and mass-based estimates of gi, whether in current-
year or older foliage. Both gi and the stomata] conductance of leaves were closely coupled to Vcmax, Jmax, and An with all variables decreasing with increasing leaf age. The N content of leaves, as well as the amount of Rubisco and other proteins, increased during the first three growing seasons, then stabilised afterwards. Thus, the age-related photosynthetic nitrogen use efficiency decline of leaves was not a consequence of decreased allocation of nitrogen towards Rubisco and other proteins. Rather, loss of photosynthetic capacity was the result of the decreased activation state of Rubisco and proportional down-regulation of electron transport towards the photosynthetic carbon reduction and photorespiratory cycles in response to a reduction of CO, supply to the chloroplasts' stroma.
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Mechanisms Controlling the Distribution of Two Invasive Bromus SpeciesBykova, Olga 15 August 2013 (has links)
In order to predict future range shifts for invasive species it is important to explore their ability to acclimate to the new environment and understand physiological and reproductive constraints controlling their distribution. My dissertation studied mechanisms by which temperature may affect the distribution of two of the most aggressive plant invaders in North America, Bromus tectorum and Bromus rubens. While Bromus tectorum is dominant in the “cold desert” steppes of the Intermountain region of western North America, B. rubens is one of the severe grass invaders in the “hot deserts” of southwestern North America. I first evaluated whether winter freezing tolerance is the mechanism responsible for the distinct northern range limits of Bromus species. Bromus rubens has a slower rate of freezing acclimation that leads to intolerance of sudden, late-autumn reductions in temperature below -12°C, Bromus tectorum, by contrast, cold hardens rapidly and is not impacted by the sudden severe late-autumn cold. Photosynthetic response to temperature does not explain their current range separation. Bromus species differ little in their photosynthetic temperature responses and the acclimation pattern of photosynthesis. Both species acclimated to a broad range of temperature through the amelioration of Pi regeneration limitation at sub-optimal temperatures and improved carboxylation capacity above the thermal optimum which probably resulted from increased thermostability of Rubisco activase. The effect of elevated temperatures during flowering on the seed yield of Bromus species demonstrates that neither species produces seed at 36°C and above. These thresholds are close to temperatures encountered during flowering in their natural environment. In summary, climatic changes will cause northward range expansion of Bromus species due to less severe autumn and winter, while reproductive failure could cause range contraction at their southern margins.
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Climate Warming and Drought Effects on Pinus and Juniperus Species: Contrasting Drought Tolerance Traits Limit Function and Growth in Tree SeedlingsLenoir, Katherine Judith 03 October 2013 (has links)
Junipers and pines exhibit contrasting patterns of growth decline and mortality with climate change-type warming and drought; yet, the underlying physiological mechanisms are not fully understood. Does warming exacerbate the effects of drought on gas exchange physiology and growth? Do the combined effects of drought and warming differ for pines and junipers? To what extent do isohydric vs. anisohydric responses to water limitation in pines and junipers constrain net leaf CO2 exchange and plant growth response to drought and warming? To address these questions, we compared responses of leaf gas exchange and growth in seedlings of juniper (Juniperus scopulorum, J. virginiana) and pine (Pinus edulis, P. taeda) species of contrasting arid and mesic origin in a study of combined warming (ambient, +1.8 °C) and enhanced summer drought (long-term mean, -40%). Warming and enhanced summer drought each reduced photosynthesis and growth and effects were largely independent, suggesting that warming exacerbates drought effects on growth. Enhanced summer drought and warming had distinct impacts on photosynthetic carbon gain that were differentially revealed depending upon soil water content. Warming reduced light-saturated net photosynthesis (Asat) under low soil water contents, whereas carry-over effects of drought treatment were evident under well-watered conditions. Short-term soil drying led to greater reduction of Asat in pines (-51%) rather than junipers (-30%). Under short-term water-limited conditions, Asat and gs were about two-times higher for junipers compared to pines. Relative growth rate of junipers declined with warming (-28%) and drought (-50%) treatments. In contrast, pine growth and Asat declined more with warming than drought. Only P. edulis exhibited increased mortality in response to warming and drought, reaching 75% in the combined warming and drought treatment. Diminished sensitivity of R to water limitations, coupled with steeper reductions in Asat with decreasing soil water content in isohydric pines compared to anisohydric junipers could account for the greater sensitivity of pines to warming and drought under climate change.
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In Search of the Holy Grail of Photoelectrochemistry : A Study of Thin Film Electrodes for Solar Hydrogen GenerationLindgren, Torbjörn January 2004 (has links)
Hydrogen is a wanted energy carrier in a future society less dependent of fossil fuels. This thesis investigates the possibilities of using solar energy to convert water into hydrogen and oxygen, so called artificial photosynthesis. Through this work multiple inexpensive and stable thin film semiconductor electrodes have been produced and used as solar energy absorbers and active sites for direct watersplitting in photoelectrochemical cells. The electrodes have mainly been of nanostructured metal oxide character but also nitrides have been studied. Detailed back ground theory on photoelectrochemistry of semiconductors for hydrogen evolution is given in the summary of the thesis. Nanostructured WO3 electrodes with a quantum yield close to unity were designed and photoelectrochemically characterized. Hematite, α-Fe2O3, nanorods were synthesized and characterized for the aim of water oxidation. The morphology of the hematite nanorods was found to be in favor of the traditional isotropic nanostructured electrodes. Moreover, a unique porous nitrogen doped TiO2 material, photoactive in visible light, was obtained by reactive sputtering. The nitrogen doped material has interesting photoelectrochemical properties and is also promising for related applications such as pollution degradation by photocatalysis. Polycrystalline indium nitride, InN, was produced by reactive sputtering. Electrodes of the as prepared InN as well as electrodes annealed in nitrogen were studied for the aim of photooxidation of water. The electrodes studied are interesting candidates as potential watersplitting electrodes in photoelectrochemical cells, even if all had in common that further improvements and optimizations need to be done.
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