Oxygen-evolving System Research Articles

An Efficient Photocatalytic Oxygen Evolution System with the Coupling of Polyoxometalates with Bismuth Vanadate

In this work, a coupling system consisting of bismuth vanadate (BiVO4) and cobalt-based polyoxometalates (Co-POMs) was developed to enhance the oxygen evolution reaction. Crystallization-driven self-assembly and the wet chemical synthesis method were deployed in synthesizing Co-POMs and monoclinic–tetragonal mixed–phase BiVO4, respectively. The introduction of Co-POMs into a BiVO4-containing mixture significantly enhanced the water oxidation reaction, with a more than twofold increment in the total amount of oxygen evolved. For instance, 461.2 µmol of oxygen was evolved from the system containing 20 mg of Co-POMs compared to 195 µmol of oxygen produced from a pristine BiVO4 system. This extraordinary improvement in the oxygen evolution reaction indicates the existence of a positive synergic effect between BiVO4 and Co-POMs, in which Co-POMs could act as effective cocatalysts to extract photogenerated charge carriers generated by BiVO4 and improve the charge transfer process. However, the amount of oxygen produced was slightly reduced to 440.7 µmol with an increase in AgNO3 loading from 30 mg to 60 mg. This unforeseen phenomenon could be elucidated by the shielding effect of silver particles, in which a higher AgNO3 loading led to a more prominent shielding effect. The presence of silver nanoparticles on post-reaction BiVO4 was confirmed by TEM and XPS analysis. This newly established process scheme provides an insight into the development of an efficient photocatalytic oxygen evolution system in realizing future commercial applications toward green energy production.

Study of the Active Sites in Porous Nickel Oxide Nanosheets by Manganese Modulation for Enhanced Oxygen Evolution Catalysis

Identifying active sites of oxygen evolution reaction catalysts is essential for studying water oxidation mechanisms. Herein, the active site of nickel oxide nanosheets by manganese modulation is investigated in an electrocatalytic oxygen evolution system. The electronic structure could be realized by Mn modulation. The intrinsic catalytic activity of Ni3+ (t2g6eg1) and Jahn–Teller active Mn3+ (t2g3eg1) species act synergistically to promote the elctrocatalytic oxygen evolution reaction. X-ray absorption near-edge structure analysis indicates that the Ni3+ and Mn3+ in Ni0.75Mn0.25 nanosheets should result from nickel vacancies and oxygen vacancies, respectively. The resulting Ni0.75Mn0.25 nanosheets show much higher oxygen evolution activity than those of NiO and Mn2O3. Density functional theory calculations indicate that the Ni and Mn act synergistically to promote formation of the O–O bond. Our work provides a comprehensive understanding of the active site of porous nickel oxide nanosheets by manganese ...

Tat transport of a Sec passenger leads to both completely translocated as well as membrane-arrested passenger proteins

We have studied the membrane transport of the chimeric precursor protein 16/33, which is composed of the Tat1-specific transport signal of OEC16 and the Sec passenger protein OEC33, both subunits of the oxygen-evolving system associated with photosystem II. Protein transport experiments performed with isolated pea thylakoids show that the 16/33 chimera is transported in a strictly Tat-dependent manner into the thylakoid vesicles yielding mature OEC33 (mOEC33) in two different topologies. One fraction accumulates in the thylakoid lumen and is thus resistant to externally added protease. A second fraction is arrested during transport in an N-in/C-out topology within the membrane. Chase experiments demonstrate that this membrane-arrested mOEC33 moiety does not represent a translocation intermediate but instead an alternative end product of the transport process. Transport arrest of mOEC33, which is embedded in the membrane with a mildly hydrophobic protein segment, requires more than 26 additional and predominantly hydrophilic residues C-terminal of the membrane-embedded segment. Furthermore, it is stimulated by mutations which potentially affect the conformation of mOEC33 suggesting that at least partial folding of the passenger protein is required for complete membrane translocation.

Hydrogen and Methanol Formation Utilizing Bioprocesses

Increases in the atmospheric levels of carbon dioxide and methane are the main causes of global warming, so measures to reduce the emissions of both gases are essential. A process converting carbon dioxide and methane into methanol was investigated combining three systems as follows, photolysis of water with solar energy, conversion of carbon dioxide and hydrogen into methane, and conversion of methane into methanol. Photolysis of water with solar energy and conversion of methane into methanol with methanotroph are described. Two different systems were attempted to achieve the photoinduced cleavage of water: photoinduced hydrogen evolution combined with Photosystem I and hydrogenase, and the combination of oxygen evolution and photoinduced hydrogen evolution systems. The oxygen evolution system contains grana and an electron carrier such as NAD and NADP. Photoinduced hydrogen evolution systems contain an electron donor, a photosensitizer, an electron carrier and a catalyst such as hydrogenase. Methane was converted into methanol by methane-oxidizing bacteria, methanotrophs which utilize methane monooxygenase to catalyze hydroxylation of methane to methanol. Using methanotrophs for methanol production, accumulation of methanol was not detected, because the methanol is subsequently oxidized by methanol dehydrogenase. The cell suspension of methanotrophs was treated by cyclopropanol as a selective inhibitor of methanol dehydrogenase, leading to extracellular methanol accumulation. The conditions of methane hydroxylation were optimized and semicontinuous methanol synthesis was achieved.

Biotic stress induced demolition of thylakoid structure and loss in photoelectron transport of chloroplasts in papaya leaves

Papaya mosaic virus (PMV) causes severe mosaic symptoms in the papaya ( Carica papaya L.) leaves. The PMV-induced alterations in photosystem II (PS II) structure and photochemical functions were probed. An increase in chlorophyll a (Chl a) fluorescence polarization suggests pathogen-induced transformation of thylakoid membrane to a gel phase. This transformation in physical state of thylakoid membrane may result in alteration in topology of pigments on pigment-binding proteins as reflected in pathogen-induced loss in the efficiency of energy transfer from carotenoids to chlorophylls. The fast Chl a fluorescence induction kinetics of healthy and PMV-infected plants by F O– F J– F I– F P transients revealed pathogen-induced perturbation on PS II acceptor side electron transfer equilibrium between Q A and Q B and in the pool size of electron transport acceptors. Pathogen-induced loss in photosynthetic pigments, changes in thylakoid structure and decrease in the ratio of F V/ F M (photochemical potential of PS II) further correlate with the loss in photoelectron transport of PS II as probed by 2,6-dichlorophenol indophenol (DCPIP)-Hill reaction. Restoration of the loss by 1,5-diphenyl carbazide (DPC), an exogenous electron donor, that donates electron directly to reaction centre II bypassing the oxygen evolving system (OES), leads towards the conclusion that OES is one of the major targets of biotic stress. Further, the data suggest that chlorophyll fluorescence could be used as a non-invasive handy tool to assess the loss in photosynthetic efficiency and symptom severity in infected green tissues vis-a-vis the healthy ones.

Prerequisites for Terminal Processing of Thylakoidal Tat Substrates

In bacteria and chloroplasts, the Tat (twin arginine translocation) system is capable of translocating folded passenger proteins across the cytoplasmic and thylakoidal membranes, respectively. Transport depends on signal peptides that are characterized by a twin pair of arginine residues. The signal peptides are generally removed after transport by specific processing peptidases, namely the leader peptidase and the thylakoidal processing peptidase. To gain insight into the prerequisites for such signal peptide removal, we mutagenized the vicinity of thylakoidal processing peptidase cleavage sites in several thylakoidal Tat substrates. Analysis of these mutants in thylakoid transport experiments showed that the amino acid composition of both the C-terminal segment of the signal peptide and the N-terminal part of the mature protein plays an important role in the maturation process. Efficient removal of the signal peptide requires the presence of charged or polar residues within at least one of those regions, whereas increased hydrophobicity impairs the process. The relative extent of this effect varies to some degree depending on the nature of the precursor protein. Unprocessed transport intermediates with fully translocated passenger proteins are found in membrane complexes of high molecular mass, which presumably represent Tat complexes, as well as free in the lipid bilayer. This seems to indicate that the Tat substrates can be laterally released from the complexes prior to processing and that membrane transport and terminal processing of Tat substrates are independent processes.

Effects of the Lack of Phosphatidylglycerol on the Donor Side of Photosystem II

Our previous studies with the pgsA mutant of the cyanobacterium Synechocystis sp. PCC6803 (hereafter termed pgsA mutant), which is defective for the biosynthesis of phosphatidylglycerol (PG), revealed an important role for PG in the electron acceptor side of photosystem II (PSII), especially in the electron transport between plastoquinones Q(A) and Q(B). This study now shows that PG also plays an important role in the electron donor side of PSII, namely, the oxygen-evolving system. Analyses of purified PSII complexes indicated that PSII from PG-depleted pgsA mutant cells sustained only approximately 50% of the oxygen-evolving activity compared to wild-type cells. Dissociation of the extrinsic proteins PsbO, PsbV, and PsbU, which are required for stabilization of the manganese (Mn) cluster, followed by the release of a Mn atom, was observed in PSII of the PG-depleted mutant cells. The released PsbO rebound to PSII when PG was added back to the PG-depleted mutant cells, even when de novo protein synthesis was inhibited. Changes in photosynthetic activity of the PG-depleted pgsA mutant cells induced by heat treatment or dark incubation resembled those of DeltapsbO, DeltapsbV, and DeltapsbU mutant cells. These results suggest that PG plays an important role in binding extrinsic proteins required for sustaining a functional Mn cluster on the donor side of PSII.

Photoinhibition in vivo of Photosystem II reactions during developmentof the photosystems of wheat seedlings

Photoinhibition of O2 evolution and reactions leading to millisecond-delayed light emission (ms-DLE) of chlorophyll by illumination of leaves with excess white light were investigated in wheat seedlings greened for different times in a special chamber with constant conditions (20 degrees C; CO2 and humidity). A sharp reduction in initial and steady state rates of O2 evolution and in the intensity of different components of ms-DLE under excess light on the stage of lag-phase of chlorophyll biosynthesis (4-6h of greening) were observed. An increasing stability of the oxygen-evolving process and ms-DLE of chlorophyll during formation of the thylakoid membrane photosystems (12-24 h of greening) was shown. Rifampicin did not influence the stability of oxygen evolution whereas cycloheximide led to the intensification of photoinhibition of the initial and steady-state rates of oxygen evolution under the inhibitory light action. The early stages of photosystems formation during short time of greening of etiolated seedlings were more sensitive to the action of inhibitory light, possibly due to a weak interaction of the oxygen-evolving system components and connection with reaction centers of Photosystem II.

Study on the photo-generation of superoxide radicals in Photosystem II with EPR spin trapping techniques.

Direct EPR evidence of the photo-generation of superoxide radicals (O(2) (-.)) was obtained by using a novel spin trapping probe in spinach Photosystem II (PS II) membrane fragments. The production of O(2) (-.) was detected by following the formation of 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) superoxide adducts (DEPMPO-OOH). The inhibition of O(2) (-.) formation by 3-(3,4-dichlorophenyl) -1,1-dimethylurea (DCMU) and the 77 K fluorescence spectrum indicated that O(2) (-.) were generated from PS II, not from PS I. The inhibition of O(2) (-.) formation by DCMU also suggested that O(2) (-.) were generated from the Q(B)binding site, not at a site prior to DCMU blockage. The extrinsic proteins and Mn are very important to eliminate O(2) (-.), showing that the oxygen-evolving system is involved in O(2) (-.) removal rather than production.

Water Oxidation Sensibilized by Tin-Porphyrin.

A tin meso-tetra(p-sulphophenyl)porphine sodium salt (SnTPPSNa4) was synthesized and its photochemical properties were characterized by measuring UV-VIS, cyclic voltammograms and fluorescence spectra. The SnTPPSNa4 was applied to a new type of photoinduced oxygen evolution system as a photosensitive oxidant and the best experimental conditions for this research are discussed in this paper. From the results, it is clear that, under the present condiation, SnTPPSNa4 could efficiently oxide water to evolve oxygen when compared to the conventional oxidant- ruthenium complex ([Ru(bpy)3] 2 + ).

Photosystem II of peas: effects of added divalent cations of Mn, Fe, Mg, and Ca on two kinetic components of P +680 reduction in Mn-depleted core particles

The catalytic Mn cluster of the photosynthetic oxygen-evolving system is oxidized via a tyrosine, Y Z, by a photooxidized chlorophyll a moiety, P + 680. The rapid reduction of P + 680 by Y Z in nanoseconds requires the intactness of an acid/base cluster around Y Z with an apparent functional p K of <5. The removal of Mn (together with bound Ca) shifts the p K of the acid/base cluster from the acid into the neutral pH range. At alkaline pH the electron transfer (ET) from Y Z to P + 680 is still rapid (<1 μs), whereas at acid pH the ET is much slower (10–100 μs) and steered by proton release. In the intermediate pH domain one observes a mix of these kinetic components (see R. Ahlbrink, M. Haumann, D. Cherepanov, O. Bögershausen, A. Mulkidjanian, W. Junge, Biochemistry 37 (1998)). The overall kinetics of P 680 + reduction by Y Z in Mn-depleted photosystem II (PS II) has been previously shown to be slowed down by divalent cations (added at >10 μM), namely: Mn 2+, Co 2+, Ni 2+, Cu 2+, Zn 2+ (C.W. Hoganson, P.A. Casey, O. Hansson, Biochim. Biophys. Acta 1057 (1991)). Using Mn-depleted PS II core particles from pea as starting material, we re-investigated this phenomenon at nanosecond resolution, aiming at the effect of divalent cations on the particular kinetic components of P + 680 reduction. To our surprise we found only the slower, proton steered component retarded by some added cations (namely Co 2+/Zn 2+>Fe 2+>Mn 2+). Neither the fast component nor the apparent p K of the acid/base cluster around Y Z was affected. Apparently, the divalent cations acted (electrostatically) on the proton release channel that connects the oxygen-evolving complex with the bulk water, but not on the ET between Y Z and P + 680, proper. Contrastingly, Ca 2+ and Mg 2+, when added at >5 mM, accelerated the slow component of P + 680 reduction by Y Z and shifted the apparent p K of Y Z from 7.4 to 6.6 and 6.7, respectively. It was evident that the binding site(s) for added Ca 2+ and Mg 2+ were close to Y Z proper. The data obtained are discussed in relation to the nature of the metal-binding sites in photosystem II.

A novel protein involved in the functional assembly of the oxygen-evolving complex of photosystem II in Synechocystis sp. PCC 6803.

Mutation of Glu69 to Gln in the D2 protein of photosystem II is known to lead to a loss of photoautotrophic growth in Synechocystis sp. PCC 6803. However, second-site mutants (pseudorevertants) with restored photoautotrophic growth but still maintaining the E69Q mutation in D2 are easily obtained. Using a genomic mapping technique involving functional complementation, the secondary mutation was mapped to slr0286 in two independent mutants. The mutations in Slr0286 were R42M or R394H. To study the function of Slr0286, mutants of E69Q and of the wild-type strain were made that lacked slr0286. Deletion of slr0286 did not affect photoautotrophic capacity in wild type but led to a marked decrease in the apparent affinity of Ca(2+) to its binding site at the water-splitting system of photosystem II and to a reduced heat tolerance of the oxygen-evolving system, particularly in E69Q. Moreover, a small increase in the half-time for photoactivation of the oxygen-evolving complex of photosystem II for both wild type and the E69Q mutant was observed in the absence of Slr0286. The accumulation of photosystem II reaction centers, dark stability of the oxygen-evolving apparatus, stability of oxygen evolution, and the kinetics of charge recombination between Q(A)(-) and the donor side were not affected by deletion of slr0286. Slr0286 lacks clear functional motifs, and no homologues are apparent in other organisms, even not in other cyanobacteria. In any case, Slr0286 appears to help the functional assembly and stability of the water-splitting system of photosystem II.

Transport of CtpA protein from the cyanobacterium Synechocystis 6803 across the thylakoid membrane in chloroplasts.

The CtpA protein in the cyanobacterium Synechocystis 6803 is a C-terminal processing protease that is essential for the assembly of the manganese cluster of the photosystem II complex. When fused to different chloroplast-targeting transit peptides, CtpA can be imported into isolated spinach chloroplasts and is subsequently translocated into the thylakoid lumen. Thylakoid transport is mediated by the cyanobacterial signal peptide which demonstrates that the protein transport machinery in thylakoid membranes is functionally conserved between chloroplasts and cyanobacteria. Transport of CtpA across spinach thylakoid membranes is affected by both nigericin and sodium azide indicating that the SecA protein and a transthylakoidal proton gradient are involved in this process. Saturation of the Sec-dependent thylakoid transport route by high concentrations of the precursor of the 33-kDa subunit of the oxygen-evolving system leads to a strongly reduced rate of thylakoid translocation of CtpA which demonstrates transport by the Sec pathway. However, thylakoid transport of CtpA is affected also by excess amounts of the 23-kDa subunit of the oxygen-evolving system, though to a lesser extent. This suggests that the cyanobacterial protein is capable of also interacing with components of the deltapH-dependent route and that transport of a protein across the thylakoid membrane may not always be restricted to a single pathway.

Parallel polarization electron paramagnetic resonance studies of the S1-state manganese cluster in the photosynthetic oxygen-evolving system.

Magnetic properties of the S1-state manganese cluster in the oxygen-evolving photosystem II were studied by parallel polarization electron paramagnetic resonance spectroscopy. Dark minus light spectra gave rise to a broad S1-state signal with a g value of about 4.9 [Dexheimer, S. L., Klein, M. P. (1992) J. Am. Chem. Soc. 114, 2821-2826]. Temperature variation of the signal intensity between 1.9 and 10 K observed in PS II with a sucrose buffer indicates that the signal originates from an excited state with a spin S of 1 with separation from the ground state (S = 0) of about 2.5 K. The S1-state signal was also observed in the sucrose buffer supplemented by 50% glycerol. However, no S1-state signal was detected by addition of 3% methanol or 30% ethylene glycol in the sucrose buffer, although illumination at 200 K in the presence of these alcohols induced the normal multiline S2 signal. Furthermore, modification of the Mn cluster by Cl- or Ca2+ depletion from PS II membranes failed to produce a detectable S1-state signal. A possible magnetic structure of the Mn cluster responsible for the generation of the S1-state signal is discussed on the basis of these observations.

S1 Destabilization and Higher Sensitivity to Light in Metribuzin-Resistant Mutants

Mutations in the secondary quinone electron acceptor (QB) pocket of the D1 protein conferring a modification on the donor side of photosystem II (PSII) have been characterized by gene cloning and sequencing in two metribuzin-resistant mutants of Synechocystis PCC 6714. The mutations induce different herbicide resistances: in M30, a point mutation at the codon 248, isoleucine to threonine, results in resistance only to metribuzin; in M35, a single mutation, Ala251Val, confers metribuzin, atrazine, and ioxynil resistance. As with other herbicide-resistant mutants, M30 and M35 present modifications in the electron transfer between the primary quinone electron acceptor (QA) and QB. In addition, they have a modified oscillatory pattern of oxygen emission: after dark adaptation, the maximum oscillation is shifted by one flash. Both mutants have a higher concentration of the redox state in the dark-adapted state than the wild type. The mutations render the oxygen-evolving system more accessible to cell reductants. The mutation Ala251Val also confers to PSII an increased sensitivity to high light. We have already demonstrated that under light stress a double mutant, AzV (Ala251Val, Phe211Ser), lost the ability to recover the PSII activity sooner than the wild type. Here, we confirm that the modification of the alanine-251 is responsible for this specific sensitivity to high light. We conclude that specific mutations of the QB pocket modify the behavior of the cells under light stress and have an effect on the structure of the D1 protein in the other side of the membrane.

Post-transcriptional and post-translational regulatory steps are crucial in controlling the appearance and stability of thylakoid polypeptides during the transition of etiolated tobacco seedlings to white light.

We have investigated the expression of nuclear-encoded chloroplast proteins that are not associated with chlorophyll (the lumenal 33-kDa and 23-kDa polypeptides of the oxygen-evolving system of photosystem II, plastocyanin and the Rieske Fe/S protein) by comparing mRNA-accumulation rates with those of the corresponding proteins during illumination of etiolated tobacco seedlings. Using subcellular fractionation, pulse/chase, Northern and Western techniques, we found that the biogenesis and stability of these proteins are regulated both translationally, as well as post-translationally, including the efficiency of mRNA uptake into polysomes, processes that operate between translation and assembly or monitor the status (soluble and membrane-attached) of a terminally processed polypeptide. Polypeptide synthesis is generally not limited by mRNA amounts. For instance, steady-state transcript levels may increase 10-fold during illumination, while those associated with polysomes increase only 2-3-fold without measurable influence on the rate of protein synthesis. The 23-kDa and Rieske polypeptides are predominantly membrane associated, but plastocyanin and the 33-kDa polypeptide are distributed among both soluble and membrane-associated protein fractions. Plastocyanin appears to be comparably stable in both forms. However, for the 33-kDa polypeptide, only the membrane-attached form is stable (> 8 h) and only this pool increases upon illumination. Its soluble form is rapidly degraded with a half-life of approximately 1 h under the chosen conditions. Our findings probably reflect part of a more general regulatory principle operating in the differentiation and maintenance of subcellular structure.

Antisense RNA for components associated with the oxygen-evolving complex and the Rieske iron/sulfur protein of the tobacco thylakoid membrane suppresses accumulation of mRNA, but not of protein.

Homologous cDNA clones of the nuclear-codes 23- and 33-kDa polypeptides of the oxygen-evolving system, as well as the Rieske iron/sulfur protein, were expressed in antisense or sense orientation under the control of the 35S RNA cauliflower mosaic virus promoter in transgenic tobacco (Nicotiana tabacum, L.) in order to modulate stationary concentrations of transcripts for studying relationships between transcript and protein levels. In all instances, in individuals that contained as little as 10% or less of the transcripts of untransformed plants, the corresponding protein levels were not notably altered. This indicated that the mRNA levels for components under study are not rate-limiting for protein accumulation and that severely reduced amounts of these transcripts still allow normal plant development. Overexpression of the 23-kDa polypeptide by the corresponding cDNA integrated in sense orientation resulted in both higher mRNA and protein levels without detectable enhancement of oxygen evolution. At least some of the excess protein was found in the soluble fraction and not associated with the photosystem-II reaction center. An attempt has been made to reconcile the varied responses obtained using these approaches with different expression patterns.

Properties of inactive Photosystem II centers

A fraction (usually in the range of 10-25%) of PS II centers is unable to transfer electrons from the primary quinone acceptor QA to the secondary acceptor QB. These centers are inactive with respect to O2 evolution since their reopening after photochemical charge separation to the S2OA (-) state involves predominantly a back reaction to S1QA in the few seconds time range (slower phases are also occurring). Several properties of these centers are analyzed by fluorescence and absorption change experiments. The initial rise phase Fo-Fpl of fluorescence induction under weak illumination reflects both the closure of inactive centers and the modulation of the fluorescence yield by the S-states of the oxygen-evolving system: We estimate typical relative amplitudes of these contributions as, respectively, 65 and 35% of the Fo-Fpl amplitude. The half-rise time of this phase is significantly shorter than for the fluorescence induction in the presence of DCMU (in which all centers are involved). This finding is shown to be consistent with inactive centers sharing the same light-harvesting antenna as normal centers, a view which is also supported by comparing the dependence of the fluorescence yield on the amount of closed active or inactive centers estimated through absorption changes. It is argued that the exponential kinetics of the Fo-Fpl phase does not indicate absence of excitation energy transfer between the antennas of inactive and active centers. We show that the acceptor dichlorobenzoquinone does not restore electron transfer in inactive centers, in disagreement with previous suggestions. We confirm, however, the enhancement of steady-state electron flow caused by this quinone and suggest that it acts by relieving a blocking step involved in the reoxidation of a fraction of the plastoquinone pool. Part of the discrepancies between the present results and those from previous literature may arise from the confusion of inactive centers characterized on a single turnover basis and PS II centers that become blocked under steady-state conditions because of deficient reoxidation of their secondary acceptors.

Manganese K-edge X-ray absorption spectra of the cyclic S-states in the photosynthetic oxygen-evolving system

A set of Mn K-edge XANES spectra due to the redox states S0-S3 of the OEC were determined by constructing a highly-sensitive X-ray detection system for use with physiologically native PS II membranes capable of cycling under a series of saturating laser-flashes. The spectra showed almost parallel upshifts with relatively high K-edge half-height energies given by 6550.9±0.2 eV, 6551.7±0.2 eV, 6552.5±0.2 eV and 6553.6±0.2 eV for the S0, S1, S2 and S3 states, respectively. The successive difference spectra between S0 and S1, S1 and S2, and S2 and S3 states were found to exhibit a similar peak around 6552-6553 eV, indicating that one Mn(III) ion or its direct ligand is univalently oxidized upon each individual S-state transition from S0 to S3. The present data, together with other observations of EPR and pre-edge XANES spectroscopy, suggest that the oxidation state of the Mn cluster undergoes a periodic change; S0: Mn(III,III,III,IV) → S1: Mn(III,IV,III,IV) → S2: Mn(III,IV,IV,IV) → S3: Mn(IV,IV,IV,IV) or Mn(III,IV,IV,IV)·L(+) with L being a direct ligand of a Mn(III) ion.

Oxygen-evolving system and secondary quinonic acceptors are highly reduced in dark adapted Euglena cells: A thermoluminescence study

Characteristics of thermoluminescence glow curves were compared in three types of Euglena cells: (i) strictly autotrophic, Cramer and Myers cells; (ii) photoheterotrophic cells sampled from an exponentially growing culture containing lactate as substrate repressing the photosynthetic activity; (iii) semiautotrophic cells, sampled when the lactate being totally exhausted, the photosynthesis was enhanced.In autotrophic and semiautotrophic cells, composite curves were observed after series of two or more actinic flashes fired at -10°C, which can be deconvoluted into a large band peaking in the range 12-22°C and a smaller one near 40°C, This second band presents the characteristics of a typical B band (due to S2/3QB (-) recombination), whereas the first one resembled the band, shifted by -15-20°C, which is observed in herbicide resistant plants. The amplitude of this major band, which was in all cases very low after one flash, exhibited oscillations of period four but rapidly damping, with maxima after two and six flashes. In contrast, photoheterotrophic Euglena displayed single, non-oscillating curves with maxima in the range 5-10°C.In autotrophic and semiautotrophic cells, oxidizing pretreatments by either a preillumination with one or more (up to twenty-five) flashes, or a far-red preillumination in the presence of methylviologen, followed by a short dark period, induced thermoluminescence bands almost single and shifted by +3-5°C, or +12°C, respectively. In autotrophic cells, far-red light plus methyl viologen treatment induced a band peaking at 31°C, as in isolated thylakoids from Euglena or higher plants, while it had barely any effect in photoheterotrophic cells.Due to metabolic activities in dark-adapted cells, a reduction of redox groups at the donor and acceptor sides of PS II dark-adapted cells is supposed to occur. Two different explanations can be proposed to explain such a shift in the position of the main band in dark-adapted autotrophic control. The first explanation would be that in these reducing conditions a decreasing value of the equilibrium constant for the reaction: SnQA (-)QB⇌SnQAQB (-), would determine the shift of the main TL band towards low temperatures, as observed in herbicide resistant material. The second explanation would be that the main band would correspond to 'peak III' already observed in vivo and assigned to S2/3QB (2-) recombinations.