1,112 research outputs found

    Origin and evolution of water oxidation before the last common ancestor of the Cyanobacteria

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    Photosystem II, the water oxidizing enzyme, altered the course of evolution by filling the atmosphere with oxygen. Here, we reconstruct the origin and evolution of water oxidation at an unprecedented level of detail by studying the phylogeny of all D1 subunits, the main protein coordinating the water oxidizing cluster (Mn4CaO5) of Photosystem II. We show that D1 exists in several forms making well-defined clades, some of which could have evolved before the origin of water oxidation and presenting many atypical characteristics. The most ancient form is found in the genome of Gloeobacter kilaueensis JS-1 and this has a C-terminus with a higher sequence identity to D2 than to any other D1. Two other groups of early evolving D1 correspond to those expressed under prolonged far-red illumination and in darkness. These atypical D1 forms are characterized by a dramatically different Mn4CaO5 binding site and a Photosystem II containing such a site may assemble an unconventional metal cluster. The first D1 forms with a full set of ligands to the Mn4CaO5 cluster are grouped with D1 proteins expressed only under low oxygen concentrations and the latest evolving form is the dominant type of D1 found in all cyanobacteria and plastids. In addition, we show that the plastid ancestor had a D1 more similar to those in early branching Synechococcus. We suggest each one of these forms of D1 originated from transitional forms at different stages towards the innovation and optimization of water oxidation before the last common ancestor of all known cyanobacteria

    Adventures with Cyanobacteria: A Personal Perspective

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    Cyanobacteria, or the blue-green algae as they used to be called until 1974, are the oldest oxygenic photosynthesizers. We summarize here adventures with them since the early 1960s. This includes studies on light absorption by cyanobacteria, excitation energy transfer at room temperature down to liquid helium temperature, fluorescence (kinetics as well as spectra) and its relationship to photosynthesis, and afterglow (or thermoluminescence) from them. Further, we summarize experiments on their two-light reaction – two-pigment system, as well as the unique role of bicarbonate (hydrogen carbonate) on the electron-acceptor side of their photosystem II, PSII. This review, in addition, includes a discussion on the regulation of changes in phycobilins (mostly in PSII) and chlorophyll a (Chl a; mostly in photosystem I, PSI) under oscillating light, on the relationship of the slow fluorescence increase (the so-called S to M rise, especially in the presence of diuron) in minute time scale with the so-called state-changes, and on the possibility of limited oxygen evolution in mixotrophic PSI (minus) mutants, up to 30 min, in the presence of glucose. We end this review with a brief discussion on the position of cyanobacteria in the evolution of photosynthetic systems

    Bioenergetic studies on the quinone electron acceptors of photosystem II

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    Photosystem II (PSII) is a membrane-bound protein complex found in plants, algae and cyanobacteria that converts light into chemical energy. Despite extensive research, many energetic and mechanistic questions of PSII remain unresolved. Here the energetics and kinetics of the electron-acceptor side of PSII from Thermosynechococcus elongatus were investigated using biophysical approaches. Based on data from electron paramagnetic resonance and thermoluminescence measurements, the two midpoint potentials of the terminal electron acceptor, QB, were measured (Em(QB/QB•−) = 92 mV; Em(QB•−/QBH2) = 43 mV). It was found that i) QB•− is significantly stabilized, contradicting the recent literature, ii) the energy-gap between QA and QB is larger than previously assumed (235 mV instead of ≈ 80 mV), contradicting the older literature, and iii) the release of QBH2 into the pool is thermodynamically favourable, ( ≈ 50 meV). No significant shift of the QB midpoint potentials in response to the loss of the Mn4O5Ca cluster was found. These findings allow for a better understanding of charge separation and the energetics of PSII. Isolated PSII from T. elongatus is used in many structural and functional studies but the electron acceptor side kinetics of this organism are poorly defined. Using absorption spectroscopy, the kinetics which were previously treated as a single “fast phase”, were resolved as follows: QA•−→ Fe 3+ (t1/2 = 50 µs); QA•−→QB(t1/2 = 350 µs); QA•−→ QB•− (t1/2 = 1.3 ms). Furthermore, the kinetic data analysis developed in this work allowed the proportions of these reactions to be determined under a range of conditions. It was found that in long dark-adapted samples up to 50% of the non-heme iron was oxidized and this oxidation was inhibited when bicarbonate was present. These data will be useful for future research on PSII and help understanding the mechanism of electron transfer on the acceptor side.Open Acces

    Dependence of substrate-water binding on protein and inorganic cofactors of photosystem II

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    The photosynthetic water oxidation reaction is catalyzed by an inorganic Mn4OxCaClyHCO3-z cluster at the heart of the oxygen evolving complex (OEC) in photosystem II. In the absence of an atomic resolution crystal structure, the precise molecular organization of the OEC remains unresolved. Accordingly, the role of the protein and inorganic cofactors of PSII (Ca2+, HCO3- and Cl-) in the mechanism of O2-evolution await clarification. In this study, rapid 18O-isotope exchange measurements were applied to monitor the substrate-water binding kinetics as a function of the intermediate S-states of the catalytic site (i.e. S3, S2 and S1) in Triton X-100 solubilized membrane preparations that are enriched in photosystem II activity and are routinely used to evaluate cofactor requirements. Consistent with the previous determinations of the 18O exchange behavior in thylakoids, the initial 18O exchange measurements of native PSII membranes at m/e = 34 (which is sensitive to the 16O18O product) show that the ‘fast’ and ‘slowly’ exchanging substrate-waters are bound to the catalytic site in the S3 state, immediately prior to O2 release. Although the slowly exchanging water is bound throughout the entire S-state cycle, the kinetics of the fast exchanging water remains too fast in the S2, S1 [and S0] states to be resolved using the current instrumentation, and left open the possibility that the second substrate-water only binds to the active site after the formation of the S3 state. Presented is the first direct evidence to show that fast exchanging water is already bound to the OEC in the S2 state. Rapid 18O-isotope exchange measurements for Ex-depleted PSII (depleted of the 17- and 23-kDa extrinsic proteins) in the S2 state reveals a resolvable fast kinetic component of 34k2 = 120 ± 14 s-1. The slowing down of the fast phase kinetics is discussed in terms of increased water permeation and the effect on the local dielectric following removal of the extrinsic subunits. In addition, the first direct evidence to show the involvement of calcium in substrate-water binding is also presented. Strontium replacement of the OEC Ca2+-site reveals a factor of ~3-4 increase in the 18O exchange of the slowly exchanging water across the S3, S2 and S1 states while the kinetics of the fast exchanging water remain unchanged. Finally, a re-investigation of the proposed role for bicarbonate as an oxidizable electron donor to photosystem II was unable to discern any 18O enrichment of the photosynthetically evolved O2 in the presence of 18O-bicarbonate. A working model for O2-evolution in terms of these results is presented

    The role of the low-molecular-weight proteins of the CP43 pre-assembly complex of Photosystem II

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    The biogenesis of Photosystem II (PS II) involves the stepwise assembly of the reaction centre subcomplex together with the subsequent addition of the two chlorophyll-binding core antenna proteins, CP43 and CP47. In this study, the function of four low-molecular-weight proteins (Psb27, Psb30, PsbK and PsbZ) belonging to the CP43 pre-assembly complex was investigated in the cyanobacterium Synechocystis sp. PCC 6803. The PsbK, PsbZ and Psb30 polypeptides are retained in the holoenzyme, but while Psb27 appears to participate in both the biogenesis and repair pathways, it is not a constituent of the mature complex. The role of conserved amino acid residues of the Psb27 protein potentially involved in protein-protein interactions with other PS II subunits was studied by introducing amino acid substitutions in Synechocystis sp. PCC 6803. These amino acid substitutions did not impede the biogenesis and repair pathways of photosystem and PS II electron transport kinetics in these mutants resembled those observed for wild type. However, chlorophyll fluorescence induction measurements indicated mutations in Psb27 altered the state transitions regulating energy transfer to the Photosystem I (PS I) and PS II and the D14A, D58E, D58K, and K63D strains exhibited changes in PBS coupling and energy transfer to PS II. Additionally, Psb27 mutations were found to alter the PS I level. Whole genome sequencing of a particular Psb27 mutant line (R54E*) revealed a C to A substitution in the psbA2 gene corresponding to a His252 to Gln substitution in the D1 reaction centre protein. Confirmation that D1:His252 was required for normal electron transfer between the primary (QA) and secondary (QB) electron acceptors was obtained by introducing point mutations into a psbA-deletion mutant of Synechocystis sp. PCC 6803. Additionally, the D1-Ser264 was also targeted, since D1-Ser264 has also been suggested to be involved in the configuration of the QB site and is in close proximity to His252. The D1-His252 mutants as well as D1-Ser264 mutants, displayed low PS II oxygen-evolving activity and impaired electron transfer between QA and QB supporting calculations suggesting His252 and Ser264 participate in the protonation of the QB- semiquinone during the turnover of the two-electron gate. Additionally, the PsbK, PsbZ and Psb30 subunits were not absolutely required for PS II activity; however, it was found that these subunits contributed to the ability of PS II to withstand photodamage following exposure to high light. Also, the Psb27 protein appears necessary for efficient repair or photoactivation of PS II and its absence along with LMW proteins (Psb30, PsbK and PsbZ) results in an increased susceptibility to photoinhibition

    Structure of a monomeric photosystem II core complex from a cyanobacterium acclimated to far-red light reveals the functions of chlorophylls d and f

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    Far-red light (FRL) photoacclimation in cyanobacteria provides a selective growth advantage for some terrestrial cyanobacteria by expanding the range of photosynthetically active radiation to include far-red/near-infrared light (700-800 nm). During this photoacclimation process, photosystem II (PSII), the water:plastoquinone photooxidoreductase involved in oxygenic photosynthesis, is modified. The resulting FRL-PSII is comprised of FRL-specific core subunits and binds chlorophyll (Chl) d and Chl f molecules in place of several of the Chl a molecules found when cells are grown in visible light. These new Chls effectively lower the energy canonically thought to define the red limit for light required to drive photochemical catalysis of water oxidation. Changes to the architecture of FRL-PSII were previously unknown, and the positions of Chl d and Chl f molecules had only been proposed from indirect evidence. Here, we describe the 2.25 angstrom resolution cryo-EM structure of a monomeric FRL-PSII core complex from Synechococcus sp. PCC 7335 cells that were acclimated to FRL. We identify one Chl d molecule in the Chl(D1) position of the electron transfer chain and four Chl f molecules in the core antenna. We also make observations that enhance our understanding of PSII biogenesis, especially on the acceptor side of the complex where a bicarbonate molecule is replaced by a glutamate side chain in the absence of the assembly factor Psb28. In conclusion, these results provide a structural basis for the lower energy limit required to drive water oxidation, which is the gateway for most solar energy utilization on earth

    Acceptor side effects on the electron transfer at cryogenic temperatures in intact photosystem II

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    AbstractIn intact PSII, both the secondary electron donor (TyrZ) and side-path electron donors (Car/ChlZ/Cytb559) can be oxidized by P680+ at cryogenic temperatures. In this paper, the effects of acceptor side, especially the redox state of the non-heme iron, on the donor side electron transfer induced by visible light at cryogenic temperatures were studied by EPR spectroscopy. We found that the formation and decay of the S1TyrZ EPR signal were independent of the treatment of K3Fe(CN)6, whereas formation and decay of the Car+/ChlZ+ EPR signal correlated with the reduction and recovery of the Fe3+ EPR signal of the non-heme iron in K3Fe(CN)6 pre-treated PSII, respectively. Based on the observed correlation between Car/ChlZ oxidation and Fe3+ reduction, the oxidation of non-heme iron by K3Fe(CN)6 at 0 °C was quantified, which showed that around 50–60% fractions of the reaction centers gave rise to the Fe3+ EPR signal. In addition, we found that the presence of phenyl-p-benzoquinone significantly enhanced the yield of TyrZ oxidation. These results indicate that the electron transfer at the donor side can be significantly modified by changes at the acceptor side, and indicate that two types of reaction centers are present in intact PSII, namely, one contains unoxidizable non-heme iron and another one contains oxidizable non-heme iron. TyrZ oxidation and side-path reaction occur separately in these two types of reaction centers, instead of competition with each other in the same reaction centers. In addition, our results show that the non-heme iron has different properties in active and inactive PSII. The oxidation of non-heme iron by K3Fe(CN)6 takes place only in inactive PSII, which implies that the Fe3+ state is probably not the intermediate species for the turnover of quinone reduction

    Discovery of a single molecule transistor in photosystem II

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    Quantum theory is used to rationalize the results of recent high-precision X-ray diffraction studies of photosystem II. It is proposed that a single molecule transistor regulates the flow of electrons through this remarkable system. At the core of the device, electrons flow through an iron(II) d-orbital by a process of superexchange, at a rate which is gated by the ambient ligand field. The transistor operates in the negative feedback mode, and its existence suggests that man-made molecular logic gates are technologically feasible. We believe this is the first recorded example of a single molecule electronic transistor in a living system

    Bicarbonate-controlled reduction of oxygen by the QA semiquinone in Photosystem II in membranes

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    Photosystem II (PSII), the water/plastoquinone photo-oxidoreductase, plays a key energy input role in the biosphere. Q∙−A, the reduced semiquinone form of the nonexchangeable quinone, is often considered capable of a side reaction with O2, forming superoxide, but this reaction has not yet been demonstrated experimentally. Here, using chlorophyll fluorescence in plant PSII membranes, we show that O2 does oxidize Q∙−A at physiological O2 concentrations with a t1/2 of 10 s. Superoxide is formed stoichiometrically, and the reaction kinetics are controlled by the accessibility of O2 to a binding site near Q∙−A, with an apparent dissociation constant of 70 ± 20 µM. Unexpectedly, Q∙−A could only reduce O2 when bicarbonate was absent from its binding site on the nonheme iron (Fe2+) and the addition of bicarbonate or formate blocked the O2-dependant decay of Q∙−A. These results, together with molecular dynamics simulations and hybrid quantum mechanics/molecular mechanics calculations, indicate that electron transfer from Q∙−A to O2 occurs when the O2 is bound to the empty bicarbonate site on Fe2+. A protective role for bicarbonate in PSII was recently reported, involving long-lived Q∙−A triggering bicarbonate dissociation from Fe2+ [Brinkert et al., Proc. Natl. Acad. Sci. U.S.A. 113, 12144–12149 (2016)]. The present findings extend this mechanism by showing that bicarbonate release allows O2 to bind to Fe2+ and to oxidize Q∙−A. This could be beneficial by oxidizing Q∙−A and by producing superoxide, a chemical signal for the overreduced state of the electron transfer chain

    The importance of the hydrophilic region of PsbL for the plastoquinone electron acceptor complex of Photosystem II

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    AbstractThe PsbL protein is a 4.5kDa subunit at the monomer–monomer interface of Photosystem II (PS II) consisting of a single membrane-spanning domain and a hydrophilic stretch of ~15 residues facing the cytosolic (or stromal) side of the photosystem. Deletion of conserved residues in the N-terminal region has been used to investigate the importance of this hydrophilic extension. Using Synechocystis sp. PCC 6803, three deletion strains: ∆(N6–N8), ∆(P11–V12) and ∆(E13–N15), have been created. The ∆(N6–N8) and ∆(P11–V12) strains remained photoautotrophic but were more susceptible to photodamage than the wild type; however, the ∆(E13–N15) cells had the most severe phenotype. The Δ(E13–N15) mutant showed decreased photoautotrophic growth, a reduced number of PS II centers, impaired oxygen evolution in the presence of PS II-specific electron acceptors, and was highly susceptible to photodamage. The decay kinetics of chlorophyll a variable fluorescence after a single turnover saturating flash and the sensitivity to low concentrations of PS II-directed herbicides in the Δ(E13–N15) strain indicate that the binding of plastoquinone to the QB-binding site had been altered such that the affinity of QB is reduced. In addition, the PS II-specific electron acceptor 2,5-dimethyl-p-benzoquinone was found to inhibit electron transfer through the quinone-acceptor complex of the ∆(E13–N15) strain. The PsbL Y20A mutant was also investigated and it exhibited increased susceptibility to photodamage and increased herbicide sensitivity. Our data suggest that the N-terminal hydrophilic region of PsbL influences forward electron transfer from QA through indirect interactions with the D–E loop of the D1 reaction center protein. Our results further indicate that disruption of interactions between the N-terminal region of PsbL and other PS II subunits or lipids destabilizes PS II dimer formation. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: Keys to Produce Clean Energy
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