The overall structure of photosynthetic pigment-protein complexes and thylakoid membranes of various photosynthetic organisms was studied using electron microscopy.

<p>This thesis deals with the impact of hydrogen bonding on the properties of phenols. The possibility for tyrosine to form hydrogen bonds to other amino acids has been found to be important for its function as an electron transfer mediator in a number of important redox enzymes. This study has focused on modeling the function of tyrosine in Photosystem II, a crucial enzyme in the photosynthetic pathway of green plants.</p><p>Hydrogen bonds between phenol and amines in both inter- and intramolecular systems have been studied with quantum chemical calculations and also in some solid-state structures involving phenol and imidazole.</p><p>Different phenols linked to amines have been synthesized and their possibilities of forming intra- and intermolecular hydrogen bonds have been studied as well as the thermodynamics and kinetics of the generation of phenoxyl radicals via oxidation reactions.</p><p>Since carboxylates may in principle act as hydrogen bond acceptors in a manner similar to imidazole, proton coupled electron transfer has also been studied for a few phenols intramolecularly hydrogen bonded to carboxylates with the aim to elucidate the mechanism for oxidation. Electron transfer in a new linked phenol—ruthenium(II)trisbipyridine complex was studied as well.</p><p>The knowledge is important for the ultimate goal of the project, which is to transform solar energy into a fuel by an artificial mimic of the natural photosynthetic apparatus</p>

Photosystem II is a unique enzyme which catalyzes light induced water oxidation. This process is driven by highly oxidizing ensemble of four Chl molecules, PD1, PD2, ChlD1 and ChlD2 called, P680. Excitation of one of the Chls in P680 leads to the primary charge separation, P680+Pheo-. Pheo- transfers electrons sequentially to the primary quinone acceptor QA and the secondary quinone acceptor QB. P680+ in turn extracts electrons from Mn4CaO5 cluster, a site for the water oxidation. There are two redox active tyrosines, TyrZ and TyrD, found in PSII. They are symmetrically located on the D1 and D2 central proteins. Only TyrZ acts as intermediate electron carrier between P680 and Mn4CaO5 cluster, while TyrD does not participate in the linear electron flow and stays oxidized under light conditions. Both tyrosines are involved in PCET. The reduced TyrD undergoes biphasic oxidation with the fast (msec-sec time range) and the slow (tens of seconds time range) kinetic phases. We assign these phases to two populations of PSII centers with proximal or distal water positions. We also suggest that the TyrD oxidation and stability is regulated by the new small lumenal protein subunit, PsbTn. The possible involvement of PsbTn protein in the proton translocation mechanism from TyrD is suggested. To assess the possible localization of primary cation in P680 the formation of the triplet state of P680 and the oxidation of TyrZ and TyrD were followed under visible and far-red light. We proposed that far-red light induces the cation formation on ChlD1. Transmembrane interaction between QB and TyrZ has been studied. The different oxidation yield of TyrZ, measured as a S1 split EPR signal was correlated to the conformational change of protein induced by the QB presence at the QB-site. The change is transferred via H-bonds to the corresponding His-residues via helix D of the D1 protein.

Doctor of Philosophy / Department of Chemistry / Ryszard J. Jankowiak / The constituents of oxygen-evolving photosystem II core complexes—antenna proteins (CP43 and CP47) and reaction center (RC)—have been the subject of many studies over the years. However, the various issues related to electronic structure, including the origin/composition of the lowest-energy traps, origin of various emission bands, excitation energy transfer (EET), primary charge separation (CS) processes and pigment site energies remain yet to be fully resolved. Exploiting our state-of-the-art techniques such as low-T absorption, fluorescence, and hole burning (HB) spectroscopies, we resolved some of the issues particularly related to CP47 and isolated RC protein complexes. For example, we demonstrated that the fluorescence origin band maximum (~695 nm) originates from the lowest-energy state ~693 nm of intact CP47. In intact CP47 in contrast to destablished protein complexes, the band (~695 nm) does not shift in the temperature range of 5–77 K unless hole-burning takes place. We also studied a large number of isolated RC preparations from spinach, and wild-type Chlamydomonas reinhardtii (at different levels of intactness), as well as its mutant (D2-L209H), in which the active branch pheophytin (PheoD1) has been genetically replaced with chlorophyll a (Chl a). We showed that the Qx-/Qy-region site-energies of PheoD1 and PheoD2 are ~545/680 nm and ~541.5/670 nm, respectively, in good agreement with our previous assignment [Jankowiak et al. J. Phys. Chem. B 2002, 106, 8803]. Finally, we demonstrated that the primary electron donor in isolated algal RCs from C. reinhardtii (referred to as RC684) is PD1 and/or PD2 of the special Chl pair (analogous to PL and PM, the special BChl pair of the bacterial RC) and not ChlD1. However, the latter can also be the primary electron donor (minor pathway) in RC684 depending on the realization of the energetic disorder. We further demonstrate that transient HB spectra in RC684 are very similar to P+QA - PQA spectra measured in PSII core, providing the first evidence that RC684 represent intact isolated RC that also possesses the secondary electron acceptor, QA. In summary, a new insight into possible charge separation pathways in isolated PSII RCs has been provided.

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2017-08-22T19:49:00Z No. of bitstreams: 1 2016 - Gepatrik Rodrigues Lima.pdf: 1199140 bytes, checksum: a162c3135ebd07b66364dfd0d3a3b7fd (MD5) / Made available in DSpace on 2017-08-22T19:49:00Z (GMT). No. of bitstreams: 1 2016 - Gepatrik Rodrigues Lima.pdf: 1199140 bytes, checksum: a162c3135ebd07b66364dfd0d3a3b7fd (MD5) Previous issue date: 2016-02-29 / The aim of the study was to evaluate the effects of herbicides applied on post- emergence on common bean cultivation, using chlorophyll a fluorescence parameters and visual evaluation, just after theirs application. The selectivity of the herbicide in the crop was also evaluated. The experiment was conducted in the crop science experimental field at the Federal Rural University of Rio de Janeiro. The treatments were the following herbicides, bentazon (720 g ha-1), fluazifop-p-butil (187,5 g ha-1), fluazifop-p-butil + fomesafen (187,5 + 250 g ha-1), fomesafen (250 g ha-1), and two other treatments with or without weeds mechanical control, and the experimental design used was on randomized blocks, with six treatments and three replications. The bentazon was the only herbicide that induced a reduction in the maximum quantum yield of PS II (Fv/Fm), in photochemical quenching (qP) and in effective quantum yield of PS II (?FSII), while it promoted an increase in non photochemical quenching (NPQ) in the first day after its application. The Fv/Fm was the best discriminatory parameter for the evaluation of the effects of the herbicide on photosynthetic apparatus of plants in the field. All the herbicides used did not promote phytotoxicity or bean yield reduction. / O trabalho teve como objetivos avaliar os efeitos da aplica??o de herbicidas p?s-emergentes na fisiologia da cultura do feijoeiro, utilizando par?metros de fluoresc?ncia de clorofila a e a seletividade do herbicida na cultura. O experimento foi conduzido no campo experimental de Fitotecnia, da Universidade Federal Rural do Rio de Janeiro. Os tratamentos constaram das aplica??es dos seguintes herbicidas: bentazon (720 g ha-1), fluazifop-p-butil (187,5 g ha-1), fluazifop-p-butil + fomesafen (187,5 + 250 g ha-1), fomesafen (250 g ha-1) e mais duas testemunhas sem aplica??o de herbicida, sendo uma capinada e outra mantida sem capina, no delineamento casualiza??o por bloco, com quatro repeti??es. Foram avaliadas as vari?veis de fluoresc?ncia da clorofila a, conte?do de prote?na sol?vel foliar e a fitotoxicidade a cultura. O bentazon foi o ?nico que causou redu??es significativas no rendimento qu?ntico m?ximo do fotossistema II (Fv/Fm), quenching fotoqu?mico (qP), rendimento qu?ntico efetivo do fotossistema II (?FSII) e promoveu incremento no quenching n?o fotoqu?mico (NPQ) nos primeiros dias ap?s sua aplica??o. O Fv/Fm foi a vari?vel mais indicada para avaliar efeitos decorrentes da aplica??o de herbicidas no aparato fotossint?tico de plantas no campo. Todos os herbicidas testados n?o ocasionaram fitotoxicidade elevada e queda no rendimento dos gr?os do feijoeiro comum.

The delivery of nutrients to the surface of marine algae can be controlled by the local hydrodynamic regime: in higher flow velocities, the Diffusive Boundary Layer (DBL) at the uptake surface is thinner, which can increase the flux of dissolved chemicals to the algal surface. If the primary productivity of an alga is controlled by the availability of a dissolved chemical, increased water flow should result in greater primary productivity due to increased chemical flux. To test the hypothesis that increased water flow will increase Photosystem II kinematics (PSII) in the green alga Caulerpa we used a Diving Pam Fluorometer to measure the maximum relative electron transport rate (Pmax), Saturation Irradiance (Ik), Non-photochemical quenching (NPQ), the light limited slope of photosynthesis vs. irradiance curve (α) and photo-chemical quenching (qP) and compared these measured values among treatments of varying flow speeds in a portable laboratory flume. We also measured the influence of water flow on values of Pmax, Ik, α , qP and NPQ in the field. Results showed that in C. racemosa collected from Tampa bay, and tested in a laboratory flume, values of Pmax and Ik were positively correlated to increase water flow, possibly indicating mass-transfer limitation. C. mexicana, collected from the Florida Keys, showed a decrease in values of Pmax, and Ik with increasing water velocity in flume experiments, indicating that the increased flow was resulting in physiological stress. This result was supported with field measurements for C. sertularioides, which showed a negative correlation between Pmax and flow velocity and Ik and flow velocity.

<p>Possibly the most fascinating biochemical mechanism remaining to be solved is the formation of oxygen from water in photosystem II. This is a critical part of the photosynthetic reaction that makes solar energy accessible to living organisms.</p><p>The present thesis uses quantum chemistry, more specifically the density functional B3LYP, to investigate a mechanism where an oxyl radical bound to manganese is the active species in O-O bond formation. Benchmark calculations on manganese systems confirm that B3LYP can be expected to give accurate results. The effect of the self-interaction error is shown to be limited. Studies of synthetic manganese complexes support the idea of a radical mechanism. A manganese complex with an oxyl radical is active in oxygen formation while manganese-oxo complexes remain inactive. Formation of the O-O bond requires a spin transition but there should be no effect on the rate. Spin transitions are also required in many short-range electron-transfer reactions.</p><p>Investigations of the superproficient enzyme orotidine decarboxylase support a mechanism that involves an invariant network of charged amino acids, acting together with at least two mobile water molecules.</p>

<p>This investigation addresses how photosynthetic efficiency is affected when pea (Pisum sativum) plants are restricted to a sole nitrogen source (i.e. ammonium or nitrate). The pea plants were watered with different nutrient solutions without NO3- or NH4+ for different time-periods in order to assay for nitrogen content. The soluble ammonium and nitrate content was measured throughout the entire growth period. No major differences were observed in nitrogen content during the starvation period up to 25 days. For technical reasons, cultivation of plants could not be extended beyond this time. The chloroplasts and thylakoids were isolated after 25 days and assayed for chlorophyll contents and photosynthetic activity.</p><p>The outcome of these tests indicates a small but unambiguous decrease in the photosynthesis activity for all treatments, relative the control.</p>

Photosystem I (PSI) from plants, algae, and cyanobacteria can mediate H2 evolution in vivo and in vitro. A simple, self-platinization procedure that permits stable PSI-mediated H2 evolution in vitro has been developed. The H2 evolution capabilities of PSI from Thermosynechococcus elongatus have been characterized. This organism utilizes cytochrome c6 (cyt c6) as the e- donor to P700. Using a solution-based, self-organized platinization of the PSI nanoparticles, this study demonstrates a sodium ascorbate-cyt-PSI-Pt-H2 electron transport and proton reduction system that yields light-dependent H2. The system was thermostable with H2 evolution increasing up to 55°C. In addition, stability studies have shown the H2 evolution to be very stable, with no significant decrease over the 80 days investigated. Through simple optimization a H2 production rate of ~5.5 mol H2/h/mg Chl [micro-mole H2 per hour per milligram chlorophyll] was attained. To further optimize the H2 production Asc-cyt-PSI-Pt-H2 system, response surface methodology (RSM) was employed. The process parameter studied included temperature, light intensity and platinum salt concentration. The results showed that experimental data had a good fit to the proposed model (R2=0.99 and p < 0.001). Platinum salt concentration, temperature and the interaction between platinum salt concentration and temperature showed significant effects on the total H2 yield. Light intensity had minimal effect of the total H2 yield within the region studied. The optimum parameters for H2 photoproduction were light intensity of 240 μE/m2/s, [micro-eistien per square meter per second], platinum salt concentration of 636 μM [micro-mol/liter] and temperature of 310C. Finally, studies that will improve the H2 yield by increasing the kinetics of electron transfer were done. A hybrid protein was formed by engineering a gene to express a fusion of the membrane-bound [Ni-Fe] hydrogenase from Ralstonia eutropha H16 and the stromal-exposed subunits PsaE and PsaD of PSI from T. elongatus. A PsaE-free mutant of PSI was simultaneously formed by genetically disrupting the expression of the PsaE subunit of a native PSI; that will allow in vitro reconstitution of the desired PsaE-hydrogenase fusion protein with PsaE-free PSI.

Photosystem II is unique! It remains the only enzyme that can oxidize water using light as energy input. Water oxidation in photosystem II is catalyzed by the CaMn4 cluster. The electrons extracted from the CaMn4 cluster are transferred to P680+ via the tyrosine residue D1-Tyr161 (YZ). Favorable oxidation of YZ is coupled to a proton transfer along a hydrogen bond to the nearby D1-His190 residue, resulting in the neutral radical YZ•. By illuminating photosystem II at cryogenic temperatures, YZ• can be trapped in a stable state. Magnetic interaction between this radical and the CaMn4 cluster gives rise to a split electron paramagnetic resonance (EPR) signal with characteristics that depend on the oxidation state (S state) of the cluster. The mechanism by which the split EPR signals are formed is different depending on the S state. In the S0 and S1 states, split signal induction proceeds via a P680+-centered mechanism, whereas in the S2 and S3 states, our results show that split induction stems from a Mn-centered mechanism. This S state-dependent pattern of split EPR signal induction can be correlated to the charge of the CaMn4 cluster in the S state in question and has prompted us to propose a general model for the induction mechanism across the different S states. At the heart of this model is the stability or otherwise of the YZ•–(D1-His190)+ pair during cryogenic illumination. The model is closely related to the sequence of electron and proton transfers from the cluster during the S cycle. Furthermore, the important hydrogen bond between YZ and D1-His190 has been investigated by following the split EPR signal formation in the different S states as a function of pH. All split EPR signals investigated decrease in intensity with a pKa of ~4-5. This pKa can be correlated to a titration event that disrupts the essential hydrogen bond, possibly by a direct protonation of D1-His190.  This has important consequences for the function of the CaMn4 cluster as this critical YZ–D1-His190 hydrogen bond steers a multitude of reactions at the cluster.