As global temperatures rise, cultivation of C3 and C4 crops in arid and semi-arid regions will face major challenges in producing biomass for billions of people. Conventional agricultural techniques that require copious irrigation will need to be complemented with dryland-farming techniques and drought-tolerant crops, such as those from the Agave genus, which use CAM photosynthesis. In the past and present, humans from arid and semi-arid regions of America have maintained a symbiotic relationship using and cultivating Agave (Agavoideae, Asparagaceae). In pre-Columbian times, Native Americans from arid regions relied on Agave cultivation as a subsistence crop to produce food, medicine, and fiber. The Hohokam in the Sonoran Desert cultivated Agave plants using rock mulching, also known as rock piles. This technique enabled the Hohokam to extensively cultivate Agave despite the limited rainwater available in the harsh Sonoran Desert. Although there are several decades of archaeological research for documenting the history of rock piles and Agave in the region beginning in the late 1970s, few studies have addressed the modern application of rock piles to cultivate Agave. Our research employed a multidisciplinary approach to bridge the historic use of rock piles to cultivate Agave with the potential application of rock piles for modern cultivation. In addition to summarizing what is known about the archaeology of Hohokam rock piles, we compiled an extensive review of the literature available on the agroecology, physiology, and natural history of Agave. We described key aspects associated with the hydrology and physical properties of Hohokam rock piles that can bolster Agave CAM photosynthesis in dry regions. We found that the use of rock piles is a feasible means of cultivating Agave under hot and dry conditions in arid regions. In addition, we used an ecological niche modeling approach and field data from Hohokam rock-pile sites and current Agave fields to assess the potential environments where rock piles could be used to cultivate Agave plants in Arizona, USA and Sonora, Mexico. We also combined an experimental archaeology approach with experimental plant physiology where we surveyed Hohokam rock-pile fields at archaeological sites to collect information about the composition of rock piles. We then created a rock-pile field where we evaluated and observed the effects of rock piles on Agave CAM utilization, mainly nocturnal CO2 uptake of Agave. Our results indicated that rock piles provide direct insulation to root systems, which indirectly benefited Agave carbon uptake and reduced temperature and drought stress. Although more agronomic research about rock pile use is needed, our research suggests that rock piles can be applied to cultivate Agave because of the physiological benefits provided such as increasing nocturnal total CO2 uptake. In addition, the suitability of rock piles in the U.S borderlands indicates that rock piles can be applied beyond the regions where they were used by the Hohokam in pre-historic times.

Water oxidation is the oxidative half-reaction of artificial photosynthesis. Realisation of effective artificial photosynthesis could present a novel method for solar energy acquisition and storage. In this thesis the first reported mononuclear ruthenium based water oxidation catalyst following second order kinetics has been theoretically investigated. Modifications to the catalyst had been reported to lead to reaction kinetics of first order. An explanation was sought by modelling and investigating multiple hypotheses. Although no conclusive explanation was found, the results indicate that the reason may be in part due to an increased propensity of the complex to undergo a competing first order rate mechanism. / Vattenoxidationsreaktionen är den oxidativa halvreaktionen i artificiell fotosyntes. Förverkligandet av artificiell fotosyntes kan leda till nya metoder för att alstra och förvara solenergi. Den här avhandlingen behandlar en teoretisk studie av den första mononukleära rutenium-baserade vattenoxidationskatalysatorn som följer andra-ordningens kinetik. Förändringar av molekylen har setts leda till en kinetik av första ordningen och en förklaring söktes genom att flertalet hypoteser testades med modellering. Någon entydig förklaring återfanns ej, men resultaten tyder på att förklaringen delvis kan ligga i en ökad benägenhet hos katalysatorn att reagera via en konkurrerande första ordningens reaktion.

>Magister Scientiae - MSc / Salt stress is a major abiotic stress that accounts for huge agricultural losses worldwide, which in turn threaten food security and sustainable agriculture. Salt triggers the excessive production of reactive oxygen species (ROS) which accumulate to levels that become toxic to plants, resulting in cell death and reduced plant growth. Part of the plant’s mechanisms to counteract ROS-induced cell death involves the scavenging ability of the antioxidant defense system to maintain redox homeostasis. Gallic acid (GA) is an antioxidant that has been shown to reduce salt-induced ROS in legume plants. However, its effects on wheat plants have not been elucidated. This study thus investigated the role of exogenous GA (250 μM) on the physiological responses and antioxidant system of wheat plants under salt stress (150 mM). In addition, this study also investigated how GA and salt stress influenced changes in the membrane proteome of wheat plants using LC-MS proteomic analysis. / 2022

Cyanobacteria are photosynthetic organisms with the potential to sustainably produce carbon-based end products by fixing carbon dioxide from the atmosphere. Optimizing the growth or biochemical production in cyanobacteria is an ongoing challenge in metabolic engineering. Rational design of metabolic pathways requires a deep understanding of regulatory mechanisms. Hence, a deeper understanding of photosynthetic regulation of the influence of the environment on metabolic fluxes provides exciting possibilities for enhancing the photosynthetic Calvin-Benson-Bassham cycle. One approach to study metabolic processes is to use omic-level techniques, such as proteomics and fluxomics, to characterize varying phenotypes that result from different environmental conditions or different genetic perturbations.<br><br>This dissertation examines the influence of light intensity on enzymatic abundances and the resulting Calvin-Benson-Bassham cycle fluxes using a combined proteomic and fluxomic approach in the model cyanobacteria Synechocystis sp. PCC 6803. The correlation between light intensity and enzymatic abundances is evaluated to determine which reactions are more regulated by enzymatic abundance. Additionally, carbon enrichment data from isotopic labelling experiments strongly suggest metabolite channeling as a flexible and light-dependent regulatory mechanism present in cyanobacteria. We propose and substantiate biological mechanisms that explains the formation of metabolite channels under specific redox conditions. <br><br>The same multi-omic approach was used to examine genetically modified cyanobacteria. Specifically, genetically engineered and conditionally growth-enhanced Synechocystis strains overexpressing the central Calvin-Benson-Bassham cycle enzymes FBP/SBPase or transketolase were evaluated. We examined the effect of the heterologous expression of each of these enzymes on the Calvin-Benson-Bassham cycle, as well as on adjacent central metabolic pathways. Using both proteomics and fluxomics, we demonstrate distinct increases in Calvin-Benson-Bassham cycle efficiency as a result of lowered oxidative pentose phosphate pathway activity. This work demonstrates the utility of a multi-omic approach in characterizing the differing phenotypes arising from environmental and genetic changes.<br><br>

This dissertation investigates the chemical ingenuity into the development of various photoactive supramolecular donor – acceptor systems to produce clean and carbon free energy for the next generation. The process is inspired by the principles learned from nature’s approach where the solar energy is converted into the chemical energy through the natural photosynthesis process. Owing to the importance and complexity of natural photosynthesis process, we have designed ideal donor-acceptor systems to investigate their light energy harvesting properties. This process involves two major steps: the first step is the absorption of light energy by antenna or donor systems to promote them to an excited electronic state. The second step involves, the transfer of excitation energy to the reaction center, which triggers an electron transfer process within the system. Based on this principle, the research is focused into the development of artificial photosynthesis systems to investigate dynamics of photo induced energy and electron transfer events. The derivatives of Porphyrins, Phthalocyanines, BODIPY, and SubPhthalocyanines etc have been widely used as the primary building blocks for designing photoactive and electroactive ensembles in this area because of their excellent and unique photophysical and photochemical properties. Meanwhile, the fullerene, mainly its readily available version C60 is typicaly used as an electron acceptor component because of its unique redox potential, symmetrical shape and low reorganization energy appropriate for improved charge separation behavior. The primary research motivation of the study is to achieve fast charge separation and slow charge recombination of the system by stabilizing the radical ion pairs which are formed from photo excitation, for maximum utility of solar energy. Besides Fullerene C60, this dissertation has also investigated the potential application of carbon nanomaterials (Carbon nanotubes and graphene) as primary building blocks for the study of the artificial photosynthesis process.

A forest’s ability to sequester carbon dioxide depends on factors such as periodic disturbance regimes, land-use change, the composition and productivity of the vegetative community, and the location and age of forested stands. However, one of the driving forces that contributes to changes in forest carbon dynamics include climatic factors, such as changes in temperature and precipitation, as well as atmospheric CO₂ concentrations which can affect the physiology of plants in complex ways. Our theorized understanding of plant physiological response to changing environmental conditions have been based on latitudinal and altitudinal studies or greenhouse experiments that measure plant physiological traits on one or a handful of plant species – and as scientists work to reduce the large variability that exists behind climate projections and plant community predictions, the need to collect locational and species-specific data becomes increasingly evident. This dissertation aims to address this issue by examining the physiological responses to temperature for 23 different tree species that have historically different geographic range distributions categorized into three groups: northern, central, and southern. The ranges of all species overlap and coexist at Black Rock Forest (BRF), an eastern deciduous forest located in the Hudson Highlands of New York. Chapter 1 examines the physiology of 16 coniferous and broadleaved tree species to determine if geographic provenance has a significant effect on foliar respiration rates, response to elevated temperature, and the respiratory substrate used to fuel the respiratory process. Chapter 2 compares the photosynthetic capacities and temperature responses of 17 broadleaved tree species to determine which range group may be more tolerant of a warming climate. Appended to this dissertation is preliminary data of a growth chamber experiment, examining the plasticity of physiological traits expressed under elevated temperatures to assess whether northern red oak seedlings show potential to acclimate to projected climate conditions and regenerate with minimal physiological constraints. Collectively, the results of these studies find significant differences in photosynthetic capacities and photosynthetic and respiration responses to temperature among species and among range groups. Northern, central, and southern ranged trees show an acclimated response to carbon assimilation under current climate conditions. However, central ranged trees, which includes the northern red oak, a dominant tree species in this region of New York, may be at a physiological disadvantage, showing lower rates of photosynthetic capacities and a trending decline of carbon assimilation under elevated temperatures. Furthermore, preliminary data from a greenhouse experiment suggests that leaf morphology and physiology traits are not plastic for northern red oak seedlings, which further weakens its physiological competitiveness and regeneration potential under warming temperatures. The results presented in this study on the physiological traits and temperature responses not only allows for a more thorough understanding of the physiological tolerances of migrant and resident tree species in the New York region but provides new data that could be incorporated into carbon and species distribution models for better predictions on carbon sequestration of forests and geographic ranges of tree species.

Molecular engineering of donor-acceptor dyads and multimodular systems to control the yield and lifetime of charge separation is one of the key goals of artificial photosynthesis for harvesting sustainably solar energy. The design of the donor-acceptor systems mimic a part of green plants and bacterial photosynthetic processes. The photochemical events in natural photosynthesis involve the capturing and funneling of solar energy by a group of well-organized chromophores referred to as an ‘antenna' system causing an electron transfer into the ‘reaction center,' where an electron transfer processes occur resulting a long-lived charge separated state. Over the last two to three decades, many efforts have been directed by the scientific community designing of multi-modular systems that are capable of capturing most of the useful sunlight and generating charge separated states of prolonged lifetimes with adequate amounts of energy. In this dissertation, we report on the design and synthesis of donor–acceptor conjugates with the goal of modulating the yield and lifetime of their charge separated states and hence, improving the conversion of light energy into chemical potential. In simple donor-acceptor systems, generally, the energy and electron transfer events originate from the singlet excited state of the donor or acceptor and can store the greatest amount of energy but must be fast to out compete intersystem crossing. To address this limitation, we have designed novel donor –acceptor conjugates that use high-energy triplet sensitizers in which electron transfer is initiated from the long lived triplet state of the donor. The triplet photosensitizers used were palladium(II) porphyrin and platinum(II) porphyrin. Heavy metal effect in these porphyrins promoted intersystem crossing and the energies of their excited state was quite high. For the case of palladium (II) porphyrin the energy stored was found to 1.89 eV and that of platinum(II) porphyrin 1.84 eV. In addition to using triplet photosensitizers as donors, we have used donors that are difficult to oxidize and hence producing long lived charge separated states with adequate amount of stored energy. The system that was used for this study is zinc porphyrin with meso-aryl pentafluorophenyl substituents and fullerene, C60 as the acceptor. The presence of fluorine substituents on zinc porphyrin makes it harder to undergo oxidation. When this high potential donor-acceptor system undergoes a photoinduced charge-separation, the estimated energy stored was found to be 1.70 eV, one of the highest reported in literature so far. To further extend the lifetime of the charge separated states generated in this high-potential zinc porphyrin-fullerene dyad a pyridine functionalized tetrathiafulvalene was axially coordinated to the Zn metal producing a supramolecular triad capable of producing long-lived charge separated state. In a subsequent study, a multi-modular donor-acceptor system composed of a porphyrin, fullerene (C60) and a BF2-chelated dipyrromethene (BODIPY) with a supramolecular arrangement in the form of porphyrin-BODIPY-C60, one of the few reported in literature. By selectively exciting BODIPY and ZnP moieties, efficient singlet-singlet energy transfer from 1BODIPY * to ZnP in toluene was observed in the case of the dyad ZnP-BODIPY. However, when ZnP is excited, electron transfer occurred with the formation ZnP.+-BODIPY-C60.- charge separated state persisting for microseconds.

Philosophiae Doctor - PhD / The tripartite symbiosis between Phaseolus vulgaris, arbuscular-mycorrhiza and the nodule bacteria, Rhizobia have been the focus of many studies ranging over a number of decades, however these studies have failed to answer certain questions relating the role of the symbionts in regard to host nutrition and the subsequent influence of these symbionts on the host C- economy. There is little doubt over the synergistic benefits involved in the dual inoculation of bean plants, as well as the resultant C-costs of maintaining the 2 symbionts, yet the specific contribution of the individual symbionts to the hosts overall nutrient and C-economy remain to be clarified. Thus the aim of this thesis is to help clarify these points by determining the symbiont induced photosynthetic, respiratory and nutritional changes taking place in the host. This was achieved by a series of experiments in which nodulated bean plants were split into two categories-those with and without AM colonized roots. These plants were then exposed to a range of growing conditions, including hi and low P, and a series of N treatments, ranging from zero N through to 3 mM NH/. Under these differing nutrient conditions growth, photosynthetic, respiratory, nutrient and amino acid responses were monitored, thus allowing for the determination of the symbionts influence on the host and the hosts reliance on the respective symbionts. Host reliance was noted most strongly under nutrient limiting conditions. Under low P treatment AM was the dominant symbiont as far as host C was concerned, allowing for the early establishment of the AM, thus ensuring the uptake of P for both host and nodule development. High P affected AM colonization to a greater extent than it did nodule dry weight and conversely the addition of N~ + led to a greater decrease in nodule dry weight than it did AM colonization. In spite of this decline, AM benefited the host by improving host N nutrition and relieving N-feedback inhibition of the export amino acid asparagine on BNF. These AM induced benefits did come at a cost to the host though, the dual inoculated plants had higher below ground respiratory costs and subsequently higher photosynthetic rates to compensate for the increased demand for C. The higher photosynthetic rates associated with dual inoculation were as a result of symbiont induced sink stimulation and not due to the improved nutrition of the host, as shown by the photosynthetic and nutrient response ratios. However, the respiratory costs associated with the uptake of soil nutrients were lower in AM colonized roots, thus showing an increased efficiency in nutrient gain by AM colonized roots. This improvement in host N nutrition as a result of AM colonization, coupled with the lower respiratory costs of AM nutrition led to the conclusion that under certain growing conditions nodules can become redundant and possibly parasitic.

<p>  </p> <p>With the increasing concern of climate change, engineering strategies to capture and fix carbon dioxide to produce valuable chemicals is a promising proposition. Metabolic engineering efforts have recently been focused on using cyanobacteria as hosts for the production biochemicals due to their ability to utilize carbon dioxide and sunlight as the sole carbon and energy sources, respectively. Unlike fermentation which uses plant derived sugars, cyanobacterial biochemical production does not compete for arable land that can be utilized for food production. Aromatic amino acids such as L-phenylalanine (Phe) and L-tryptophan (Trp) are essential amino acids since they cannot be synthesized by animals and thus are needed as supplements. They are valuable as animal feed supplements in the agricultural industry and find wide applications in the food, cosmetic and pharmaceutical industries as precursors. However, investigation of cyanobacteria for production of aromatic amino acids such as Phe and Trp is limited. This dissertation studies (<em>i</em>) combining random mutagenesis and metabolic engineering techniques for Trp and Phe production in <em>Synechocystis </em>sp. PCC 6803, (<em>ii</em>) development of a fast-growing cyanobacteria strain <em>Synechococcus elongatus</em> PCC 11801 for Phe production and (<em>iii</em>) investigating the effect of creation of Phe sink on photosynthetic efficiency under different light intensities.</p> <p>Aromatic amino acid biosynthesis is tightly regulated by feedback inhibition in cyanobacteria. To enable overproduction of Trp in <em>Synechocystis</em> sp PCC 6803, we utilized chemical mutagenesis coupled with analog selection followed by genome sequencing to identify single nucleotide polymorphisms (SNPs) responsible for the Trp overproduction phenotypes. Interestingly, overproducers had mutations in the competing Phe biosynthetic pathway gene chorismate mutase (CM) which resulted in a lower enzyme activity and redirection of flux to Trp. We subsequently overexpressed genes encoding feedback insensitive enzymes in our randomly engineered Trp overproducing strain. The best strain isolated was able to accumulate 212±23 mg/L Trp in 10 days under 3% (vol/vol) CO2. We demonstrate that combining random mutagenesis and metabolic engineering is superior to either approach alone.</p> <p>Initial efforts in engineering cyanobacteria have resulted in low titers and productivities due to slow growth. Recently a fast-growing cyanobacterial strain <em>Synechococcus elongatus</em> PCC 11801 was discovered with growth rates comparable to yeast. Due to the lack of well characterized synthetic biology tools available for metabolic engineering of this strain, we use two rounds of ultraviolet (UV) mutagenesis and analog selection to develop Phe overproducing strains. The best strain obtained using this strategy can produce 1.2 ± 0.1 g/L of Phe in 3 days under 3% (vol/vol) CO2. This is the highest titer and productivity for Phe production currently reported by cyanobacteria highlighting the promise of engineering fast-growing strains for biochemical production.</p> <p>Interestingly, Phe overproduction does not compete with growth but happens by fixing carbon at a higher rate. It is thought that the introduction of this carbon and energy sink relieves “sink limitation” by improving light use. However, neither the molecular mechanism nor the effect of light on enhancement in carbon fixation by introduction of an additional sink are known. Therefore, we investigated the effect of light intensity on photosynthetic efficiency, linear and cyclic electron flow in the strain containing the Phe sink. Our results indicate that under excess light, introduction of the Phe sink improves carbon fixation by improving photosynthetic efficiency and substantially reducing the cyclic electron flow around photosystem I (PSI). Taken together, our results show the previously untapped potential of cyanobacteria to improve carbon fixation by the unintuitive strategy of introducing a native carbon product sink and highlight the importance of the light environment on its performance.</p> <p>Although further improvements in titer, productivity, and scale up will be necessary for cyanobacteria to compete economically at the industrial scale, this dissertation adds to the scientific knowledge and techniques for further metabolic engineering efforts.</p>

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