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... The release of protons has been observed in response to changes in the oxidation state (the S n state, where the subscript represents the number of oxidation steps accumulated) of the oxygen-evolving complex and occurs with a typical stoichiometry of 1:0:1:2 for the S 0 -S 1 -S 2 -S 3 -S 0 transitions, respectively (for example, refs 3,4). Although the relevant pathway for proton transfer (PT) in each S-state transition is not yet clear, PT may proceed via different pathways in the PSII protein, depending on the S-state transition [5][6][7] . Candidates for the relevant PT pathways have been reviewed recently [7][8][9][10][11][12] and site-directed mutagenesis studies are testing the various possibilities 13 . ...
... Although the relevant pathway for proton transfer (PT) in each S-state transition is not yet clear, PT may proceed via different pathways in the PSII protein, depending on the S-state transition [5][6][7] . Candidates for the relevant PT pathways have been reviewed recently [7][8][9][10][11][12] and site-directed mutagenesis studies are testing the various possibilities 13 . ...
... 59). PT may proceed through different pathways depending on the S-state transitions [5][6][7] . Intriguingly, it has been reported that the rate constant for the S 0 -S 1 transition is essentially pH-independent, whereas that for S 2 -S 3 transition is pH-dependent 21,48,49 . ...
In photosystem II (PSII), the Mn 4 CaO 5 cluster catalyses the water splitting reaction. The crystal structure of PSII shows the presence of a hydrogen-bonded water molecule directly linked to O4. Here we show the detailed properties of the H-bonds associated with the Mn4CaO5 cluster using a quantum mechanical/molecular mechanical approach. When O4 is taken as a μ-hydroxo bridge acting as a hydrogen-bond donor to water539 (W539), the S0 redox state best describes the unusually short O4-O W539 distance (2.5 Å) seen in the crystal structure. We find that in S1, O4 easily releases the proton into a chain of eight strongly hydrogen-bonded water molecules. The corresponding hydrogen-bond network is absent for O5 in S1. The present study suggests that the O4-water chain could facilitate the initial deprotonation event in PSII. This unexpected insight is likely to be of real relevance to mechanistic models for water oxidation.
... Besides its physiological role (see above), Cyanobacteria and soil algae need Cl − for splitting water in oxygenic photosynthesis (Renger, 2001). In bound form, Cl appears in many organochlorines. ...
... Beside heterocyst formation, Ca 2+ appears in having a general role in N 2 -fixation by Cyanobacteria (Gallon, 1992). At photosystem II of Cyanobacteria and algae, Ca is involved in water splitting (Renger, 2001;Umena et al., 2011). ...
The soil microbial community fulfils various functions, such as nutrient cycling and carbon (C) sequestration, therefore contributing to maintenance of soil fertility and mitigation of global warming. In this context, a major focus of research has been on C, nitrogen (N) and phosphorus (P) cycling. However, from aquatic and other environments, it is well known that other elements beyond C:N:P are essential for microbial functioning. Nonetheless, for soil microorganisms this knowledge has not yet been synthesised. To gain a better mechanistic understanding of microbial processes in soil systems, we aimed at summarising the current knowledge on the function of a range of essential or beneficial elements, which may affect the efficiency of microbial processes in soil. This knowledge is discussed in the context of microbial driven nutrient and C cycling. Our findings may support future investigations and data evaluation, where other elements than C:N:P affect microbial processes.
... Lastly, we emphasize that in this new proposed mechanism, the essential charge rearrangement involved in the steps of S 3 -Y Z • → S 4 → S 0 is consistent with the kinetic studies where a slow kinetic phase involving a structural rearrangement has been strongly suggested, 19,[21][22][23]27,28 giving a possible explanation to this slow kinetic phase. Two molecules of water are transferred into the Mn 4 CaO 5 cluster during each catalytic cycle: one from the Ca water channel (as the successor of O5 for the next catalytic cycle) and one from the Asp61 water channel (for the new W2) consumed for forming O 2 . ...
... tively charged environment around the Mn 4 CaO 5 cluster and the pK a balance of proton transfer channels.20,27,28 These multiple processes involved in the S 3 → S4 step, including storage of one charge on Y Z • , release of one proton, charge rearrangement, and formation of the Mn VII -dioxo site, are crucial for restoring the severely charged Mn cluster and producing a standby state to accept a fourth charge and form the O-O bond. ...
Resolving the questions, namely, the selection of Mn by nature to build the oxygen-evolving complex (OEC) and the presence of a cubic Mn3CaO4 structure in OEC coupled with an additional dangling Mn (Mn4) via μ-O atom are not only important to uncover the secret of water oxidation in nature, but also essential to achieve a blueprint for developing advanced water-oxidation catalysts for artificial photosynthesis. Based on the important experimental results reported so far in the literature and on our own findings, we propose a new hypothesis for the water oxidation mechanism in OEC. In this new hypothesis, we propose for the first time, a complete catalytic cycle involving a charge-rearrangement-induced MnVII–dioxo species on the dangling Mn4 during the S3 → S4 transition. Moreover, the O–O bond is formed within this MnVII–dioxo site, which is totally different from that discussed in other existing proposals.
... The possibility of O-O (or O-OH) bond formation in the S 3 state has been a classic problem extensively investigated by Renger [104] since electron-transfer (ET) dynamics experiments by his group [103] have indicated the maximum activation barrier in the S 3 state (see sup- porting section SIV) [107]. Our previous QM computa- tions [99,105,106] also indicated that several interme- diates with different ground spin states are feasible in the S 3 state. ...
... Here, AB-type mechanisms for the O-O bond formation are revisited on the theoretical grounds. Several groups [68][69][70][71][72]100,104,107] already proposed initial formation of the O-OH bond for oxygen evolution in OEC of PSII instead of the O-O bond formation in Section 5. Berkeley group [51] discussed possible mechanisms of formation of the O-OH bond in the S 4 state on the basis of the SFX results. Their proposals are consistent with our previous models in Scheme 5 [33]. ...
Possible mechanisms for water cleavage in oxygen evolving complex (OEC) of photosystem II (PSII) have been investigated based on broken-symmetry (BS) hybrid DFT (HDFT)/def2 TZVP calculations in combination with available XRD, XFEL, EXAFS, XES and EPR results. The BS HDFT and the experimental results have provided basic concepts for understanding of chemical bonds of the CaMn4O5 cluster in the catalytic site of OEC of PSII for elucidation of the mechanism of photosynthetic water cleavage. Scope and applicability of the hybrid DFT (HDFT) methods have been examined in relation to relative stabilities of possible nine intermediates such as Mn-hydroxide, Mn-oxo, Mn-peroxo, Mn-superoxo, etc., in order to understand the O–O (O–OH) bond formation in the S3 and/or S4 states of OEC of PSII. The relative stabilities among these intermediates are variable, depending on the weight of the Hartree–Fock exchange term of HDFT. The Mn-hydroxide, Mn-oxo and Mn-superoxo intermediates are found to be preferable in the weak, intermediate and strong electron correlation regimes, respectively. Recent different serial femtosecond X-ray (SFX) results in the S3 state are investigated based on the proposed basic concepts under the assumption of different water-insertion steps for water cleavage in the Kok cycle. The observation of water insertion in the S3 state is compatible with previous large-scale QM/MM results and previous theoretical proposal for the chemical equilibrium mechanism in the S3 state . On the other hand, the no detection of water insertion in the S3 state based on other SFX results is consistent with previous proposal of the O–OH (or O–O) bond formation in the S4 state . Radical coupling and non-adiabatic one-electron transfer (NA-OET) mechanisms for the OO-bond formation are examined using the energy diagrams by QM calculations and by QM(UB3LYP)/MM calculations . Possible reaction pathways for the O–O and O–OH bond formations are also investigated based on two water-inlet pathways for oxygen evolution in OEC of PSII. Future perspectives are discussed in relation to post HDFT calculations of the energy diagrams for elucidation of the mechanism of water oxidation in OEC of PSII.
... Th e formation of photosynthetic molecular oxygen takes place within the multiple enzymatic water oxidizing complex (WOC) of photosystem II (PS II) of plants and cyanobacteria as a result of four-electron oxidation of two water molecules (see Renger, 2001). Th e basis of the WOC is a so-called inorganic core (Mn-cluster) composed of four atoms of Mn, one atom of Ca and fi ve atoms of O (Umena et al., 2011). ...
... Conditionally, PS II can be divided into two main blocks: (1) the photochemical RC in which energy of the excited chlorophyll absorbing a photon of light converts to energy of the separated states resulting in the formation of the very strong biological oxidant -chlorophyll P 680 + (with the redox potential equal to 1.10 -1.27 V (Klimov et al., 1979;Rappaport et al., 2002;Allakhverdiev et al., 2010)); and (2) the water-oxidizing complex which is repeatedly oxidized by the P 680 + through the secondary electron donor, Tyr Z , and in turn oxidizes H 2 O to O 2 (see Renger, 2001). ...
In recent years significant progress in the study of the structural and functional organization of the Mn-cluster (inorganic core) of the water-oxidizing complex (WOC) of photosystem II (PS II) has been achieved. Despite this fact, the question about the evolutionary origin of the inorganic core of the WOC of PS II still remains open. The results of electrochemical and EPR measurements show that in the presence of bicarbonate ions, the oxidation of Mn2+ cations is significantly facilitated upon formation of Mn²⁺-bicarbonate complexes. The oxidation potential of Mn2+ to Mn3+ in the Mn²⁺-bicarbonate complex is low enough, that the photooxidation of Mn²⁺ by reaction centers (RCs) of anoxygenic photosynthetic bacteria could be expected. Based on this, an assumption about a possible role of the "low-potential" Mn²⁺-bicarbonate complexes in the evolutionary origin of the Mn-cluster of the WOC of PS II was made. Such complexes could be used by anoxygenic bacteria containing type II RCs initially as electron donors, and then as "building blocks" for the formation of the enzymatic Mn-containing center capable to do the water oxidation that could lead to the appearance of the first O2-evolving cyanobacteria. This chapter describes current research in this area.
... where P680 is the photoactive chlorophyll-a (Chl-a) component, Pheo is the primary acceptor, k PC is the rate constant of primary radical pair formation from P680 + and Q A is a noncovalently bound plastoquinone-9 molecule (PQ-9) [19,20]. This reaction is kinetically limited by N stab that was found to be about (300ps) -1 [21]. ...
... The S -2 EPR multiline signal can be very well simulated by a magnetically isolated Mn 2 (II, III) dimer of spin S = 1/2[161]. For unknown reasons the other two Mn ions of the OEC do not contribute to this signal, but based on the above assignment for the S 1 state the redox states of the whole cluster should be Mn 4 (II, III, III, III) in the S -2 state.In light of the emerging details about the structure of PS II in the thermophilic cyanobacterium 7 HORQJDWXV based on X-ray crystallography[25,26,261] and the lack of crystallographic data about the otherwise better studied PS II from higher plants[20,164,262,263] it is interesting to compare the structure and function of the two systems in detail. In this regard EPR measurements can play an important role because magnetic couplings are sensitive to changes in the structure and ligands of the OEC. ...
In Cyanobakterien und Pflanzen erfolgt die Spaltung von Wasser in molekularen Sauerstoff, vier Protonen und vier Elektronen in einem Pigment-Protein Komplex, der Teil der Thylakoidmembran ist und als Photosystem II (PS II) bezeichnet wird. Die Wasserspaltung wird von einem funktionellen Teil des PS II katalysiert, der als Sauerstoff entwickelnder Komplex ( Oxygen Evolving Complex , OEC) bekannt ist und dessen katalytisches Zentrum von einem Mn4OxCa-Komplex gebildet wird. Der OEC durchläuft während der Wasseroxidation fünf Redoxzustände (S-Zustände). Die kürzlich veröffentlichten Kristallstrukturen von PS II aus den thermophilen Cyanobakterien Thermosynechococcus elongatus (T. elongatus) und T. vulcanus liefern Informationen über die komplexe Gesamtstruktur von PS II und die Anordnung vieler Kofaktoren. Bis heute existiert keine Kristallstruktur für PS II aus höherer Pflanzen. Da bisher die meisten funktionellen Untersuchungen an PS II Komplexen aus Spinat durchgeführt wurden, sind für eingehende Untersuchungen von Struktur-Funktionsbeziehungen vergleichende Funktionsstudien an PS II Komplexen aus Cyanobakterien und höheren Pflanzen erforderlich. Ziel dieser Arbeit ist die Untersuchung von Gemeinsamkeiten und Unterschieden beim Mechanismus der Sauerstoff-Entwicklung in T. elongatus und Spinat. Hierfür wurden insbesondere blitzinduzierte Sauerstoff-Oszillationsmuster ( Flash Induced Oxygen evolution Patterns , FIOPs) unter verschiedensten Bedingungen gemessen und im Rahmen eines erweiterten Kok-Modells analysiert. Es wurden folgende Ergebnisse erzielt: a) Die Temperatur-Abhängigkeiten der miss - und double hit -Wahrscheinlichkeiten, sowie die Lebensdauern der S-Zustände in beiden Organismen deuten auf strukturelle Unterschiede der Akzeptorseite von PS II und in der Umgebung des Tyrosin-Radikals YDox hin. b) Untersuchungen zu den Effekten von einem H/D Isotopen-Austausch auf die Reaktionen des OEC bei verschiedenen Temperaturen und pL-Werten (L = H oder D) zeigen, dass hierdurch die Reaktionen im PS II beider Organismen in vergleichbarer Weise beeinflusst werden. Im Gegensatz zu Spinat-Thylakoiden ist in Thylakoiden von T. elongatus aber YDox, das bei pH 7,0 stabil ist, bei pH 8,0 labil und wird im Dunkeln bei Raumtemperatur innerhalb einer Stunde reduziert. c) Durch Inkubation mit den exogenen Reduktionsmitteln NH2OH, N2H4 und NO wurde zum ersten Mal gezeigt, dass (i) Arrhenius-Diagramme für NH2OH induzierte S1 ? S0 and S0 ? S-1 Übergänge im OEC von Spinat-Thylakoiden einen Knickpunkt bei 29°C aufweisen. Unterhalb dieser Temperatur sind Aktivierungsenergie und prä-exonentielle Faktoren unabhängig von S-Zustand, wogegen oberhalb von 29°C beide Faktoren vom Redoxzustand abhängig sind. (ii) Das S-2 EPR Multiline -Signal wurde erstmals in monomeren und dimeren PS II core -Komplexen von T. elongatus durch NO -Inkubation erhalten. Kleine, reproduzierbare Verschiebungen einiger Tieffeld-Übergänge in den S-2 EPR- Multiline -Signalen von T. elongatus im Vergleich zu dem Spinat-Signal weisen auf leichte Unterschiede bei der Koordinations-Geometrie und/oder den Liganden des Mn4OxCa-Komplexes zwischen thermophilen Cyanobakterien und höheren Pflanzen hin. (iii) FIOPs von N2H4-reduzierten Thylakoiden von T. elongatus zeigen eine Besetzung des S-3 -Zustandes von bis zu ~70% an; die Stabilität dieses Redoxzustandes macht die Existenz eines Mn4(II4)-Komplexes für diesen Zustand unwahrscheinlich. Darüber hinaus weist die numerische Analyse der FIOPs von mit Hydrazin reduziertem PS II auf die Existenz von S-4- und S-5- Redoxzuständen hin. Diese Ergebnisse unterstützen die Zuordnung der Mangan-Oxidationszustände Mn4(III2,IV2) für den S1-Zustand.
... The water oxidising complex of photosystem 2 oxidises two water molecules to molecular oxygen at a rate approaching 1000 s -1 at ambient temperatures and pressure. [1][2][3] It is one of the most important reactions in biology, and is also of intense interest from a green energy perspective, where it is recognized to be the main barrier to the development of commercial solar devices for the generation of hydrogen from water. 4 Water oxidation to dioxygen is a challenging due to the high endergonicity (E° = 0.82 V (vs NHE) at pH 7) for the reaction, 5 Somewhat similar proposals have been put forward for the WOC namely water nucleophilic attack 6 or oxyl radical-oxo coupling 7 These require the generation of a reactive oxo species in the final Kok-cycle S 4 state. Artificial catalysts generally use very high strength oxidising agents to generate reactive oxo species, either radical oxygen species or highly charged metal electrophilic species. ...
In this report we combine broken symmetry density functional calculations and electron paramagnetic resonance analysis to obtain the electronic structure of the penultimate S3 state of Nature’s water oxidising complex and determine the electronic pathway of O-O bond formation. Analysis of the electronic structure changes along the reaction path shows that two spin crossovers, facilitated by the geometry and magnetism of the water oxidising complex are used to provide a unique low energy pathway. The pathway is facilitated via formation and stabilisation of the [O2]3- ion. This ion is formed between ligated deprotonated substrate waters, O5 and O6, and is stabilised by antiferromagnetic interaction with the Mn ions of the complex. Combining computational, crystallographic and spectroscopic data we show that an equilibrium exists between an O5 oxo and O6 hydroxo form with an S=3 spin state and a deprotonated O6 form containing a two-centre one electron bond in [O5O6]3- which we identify as the form detected using XFEL crystallography. This form gives rise to an S=6 spin state which we demonstrate gives rise to a low intensity EPR spectrum compared with the accompanying S=3 state, making its detection via EPR difficult and overshadowed by the S=3 form. Simulations using 50% -70% of the S=6 component give rise to a superior fit to the experimental W- band EPR spectral envelope compared with an S=3 only form. The computational , crystallographic and spectroscopic data are shown to coalesce to the same picture of a predominant S=6 species containing the first one-electron oxidation product of two water molecules i.e. [O5O6]3-. Progression of this form to the two-electron oxidised peroxo and three-electron oxidised superoxo forms, leading eventually to the evolution of triplet O2 , is shown to be the pathway Nature adopts to oxidise water under ambient conditions. The study reveals the key electronic, magnetic and structural design features of Nature’s catalyst which facilitates water oxidation to O2 under ambient conditions.
... Thus, the open-closed isomerization in the S 3 n Y z • state may correspond to the proposed "structural isomerization" preceding dioxygen formation 93 and to thereby constitute the rate limiting 1−2 ms phase (slow phase) that follows a 200 μs lag phase ( Figure 1d) and precedes the much more rapid O 2 formation. 3, [12][13][14][15][16][17][18][19][20][21][22]92,127 According to the Eyring−Polanyi equation of TS theory assuming a standard pre-exponential factor, 128−130 the 1−2 ms kinetics is calculated to correspond to an activation free energy ∼14 kcal/mol at room temperature. Given the limited errors from DFT methodology and possibly experimental measurement, a safer quantity for the barrier should be around 13−15 kcal/mol for a process that occurs on a timescale of milliseconds. ...
Photosynthetic water oxidation is catalyzed by a manganese-calcium oxide cluster, which experiences five "S-states" during a light-driven reaction cycle. The unique "distorted chair"-like geometry of the Mn4CaO5(6) cluster shows structural flexibility that has been frequently proposed to involve "open" and "closed"-cubane forms from the S1 to S3 states. The isomers are interconvertible in the S1 and S2 states, while in the S3 state, the open-cubane structure is observed to dominate inThermosynechococcus elongatus (cyanobacteria) samples. In this work, using density functional theory calculations, we go beyond the S3+Yz state to the S3nYz• → S4+Yz step, and report for the first time that the reversible isomerism, which is suppressed in the S3+Yz state, is fully recovered in the ensuing S3nYz• state due to the proton release from a manganese-bound water ligand. The altered coordination strength of the manganese-ligand facilitates formation of the closed-cubane form, in a dynamic equilibrium with the open-cubane form. This tautomerism immediately preceding dioxygen formation may constitute the rate limiting step for O2 formation, and exert a significant influence on the water oxidation mechanism in photosystem II.
... The more increase in canopy temperature of A. indica as compared to T. mantaly during the hotter period of the day might be due to occurrence of less transpiration under drought stress (Turner et al. 2001) and capacity of plant to decrease in leaf area in response to drought stress just to cut down the water budget at the cost of yield loss (Schuppler et al. 1998). Photosystem II (PSII) is a light-dependent water:plastoquinone-oxidoreductase that uses light energy to oxidize water and to reduce plastoquinone (Renger 2001). As canopy temperature is an indicator of plant water status (Amani et al. 1996) and drought might cause damage to the oxygen-evolving center (OEC) coupled with PSII (Kawakami et al. 2009) due to less available tissue water, the resulted higher capacity to maintain PSII efficiency in A. indica as compared to T. mantaly during desiccation might be due to low vulnerability to chronic photo inhibition and photo inactivation of photosystem II centers. ...
... The more increase in canopy temperature of A. indica as compared to T. mantaly during the hotter period of the day might be due to occurrence of less transpiration under drought stress (Turner et al. 2001) and capacity of plant to decrease in leaf area in response to drought stress just to cut down the water budget at the cost of yield loss (Schuppler et al. 1998). Photosystem II (PSII) is a light-dependent water:plastoquinone-oxidoreductase that uses light energy to oxidize water and to reduce plastoquinone (Renger 2001). As canopy temperature is an indicator of plant water status (Amani et al. 1996) and drought might cause damage to the oxygen-evolving center (OEC) coupled with PSII (Kawakami et al. 2009) due to less available tissue water, the resulted higher capacity to maintain PSII efficiency in A. indica as compared to T. mantaly during desiccation might be due to low vulnerability to chronic photo inhibition and photo inactivation of photosystem II centers. ...
Infrared (IR) imaging, chlorophyll fluorescence imaging and plant phenomic approach were used to study physiological mechanism of desiccation tolerance in Azadirachta indica and Terminalia mantaly during the period of November 2018 to February 2019. IR imaging instrument was installed in the field for monitoring the canopy temperature dynamics of different canopy level including stem region of the tree throughout the day. Maximum photochemical efficiency (Fv/Fm) was measured with chlorophyll fluorescence measuring system for sun exposed leaves of A. indica and T. mantaly over a period of desiccation. In order to reveal complete understanding of physiological mechanism of desiccation tolerance, plant phenomic approach was used for assessing response of these tree species to exposed desiccation. Results indicated that canopy temperature of upper foliage, lower foliage, stem (trunk) region of A. indica were quite higher during the hotter period of the day as compared to T. mantaly and maximum photochemical efficiency (Fv/Fm) was maintained in A. indica leaves as compared to T. mantaly for same exposed duration of desiccation. Plant phenomic approach also depicted that A. indica twig retained more tissue water and maintained canopy volume area higher than T. mantaly. Thus it provides an indication that A. indica tree is quite desiccation tolerant than T. mantaly by maintaining its canopy temperature, maximum photochemical efficiency, more tissue water and canopy area.
... Green plants use carbon dioxide (CO 2 ), water and the light energy to synthesize sugars (Nelson, 2011), releasing O 2 as a 'waste' product. The O 2 released can then act as a substrate for many aspects of cellular metabolism (Renger, 2001). More than 350 O 2 -dependent reactions have been reported to occur in the various metabolic pathways found in living organisms (Raymond and Segrè, 2006;Schmidt et al., 2018). ...
Molecular oxygen (O 2 ) is a basic requirement of life for many organisms, including plants. As a terminal electron acceptor, during mitochondrial respiration, O 2 governs both the energy status and numerous aspects of cellular metabolism by influencing adenosine triphosphate (ATP) synthesis. Although O 2 is essential, no active mechanism has been found in plants that can provide uniform O 2 within the tissues of various plant organs, including roots and tubers, as well as germinating seeds. As a result, plants cells and tissues can face low-oxygen stress (LOS)/hypoxia under certain environmental conditions. Environmental events such as flooding/water logging can create LOS or absence of O 2 , and a high rate of cellular metabolism can also cause the O 2 deficit. The absence of O 2 as a terminal electron acceptor leads to the generation of excess electrons that can leak from the inner mitochondrial membrane, resulting in the production of reactive oxygen species (ROS). With respect to physiological signaling, ROS and nitric oxide (NO) have shown to have a dual behaviour, depending upon the cellular concentrations and/or time of exposure. Both ROS and NO can act as signaling molecules, and depending on their endogenous levels they can activate various downstream signaling pathways or can cause an oxidative burst, leading to cell damage. In addition, previous studies have shown that ROS and NO, when being present at high levels, can react with each other to produce various other forms of ROS and reactive nitrogen species (RNS). The present review focuses on different aspects of LOS and how plants sense decreases in cellular O 2 concentrations. The roles played by ROS and NO in relation to maintaining redox homeostasis during impairment of energy metabolism are critically discussed.
... During the photosynthetic light reaction, plants and algae release oxygen (O 2 ) into the atmosphere as a by-product of water oxidation [1]. Photosynthetic water oxidation takes place in photosystem II (PSII), which is present in the thylakoid membrane and functions as a waterplastoquinone oxidoreductase [2]. Photosynthetic oxygen evolution requires only moderate activation energies, and PSII can release up to 50 O 2 molecules per second [1,3]. ...
The excessive and harmful light energy absorbed by the photosystem (PS) II of higher plants is dissipated as heat through a protective mechanism termed non-photochemical quenching (NPQ) of chlorophyll fluorescence. PsbSknock-out (KO) mutants lack the trans-thylakoid proton gradient (ΔpH)-dependent part of NPQ. To elucidate the molecular mechanism of NPQ, we investigated its dependency on oxygen. The development of NPQ in wild-type (WT) rice under low-oxygen (LO) conditions was reduced to more than 50% of its original value. However, under high-oxygen (HO) conditions, the NPQ of both WT and PsbS-KO mutants recovered. Moreover, WT and PsbS-KO
mutant leaves infiltrated with the ΔpH dissipating uncoupler nigericin showed increased NPQ values under HO conditions. The experiments using intact chloroplasts and protoplasts of Arabidopsis thaliana supported that the LO effects observed in rice leaves were not due to carbon dioxide deficiency. There was a noticeable 90% reduction in the half-time of P700 oxidation rate in LO-treated leaves compared with that of WT control leaves, but the HO treatment did not significantly change the half-time of P700 oxidation rate. Overall, the results obtained here indicate that the stroma of the PsbS-KO plants could be potentially under O2 deficiency. Because the functions of PsbS in rice leaves are likely to be similar to those in other higher plants, our findings offer novel insights into the role of oxygen in the development of NPQ.
... For developing an artificial system with efficiency close to the natural photosynthesis, in-depth knowledge about the oxygen evolution mechanism in nature is critical. Although several potential models have been proposed (Renger, 2001;Shen, 2015), the exact mechanism is still not fully understood. A computational study showed that two water molecules bound to calcium and the fourth manganese ion play a key role in the water-splitting process (Barber, 2016) and that the three other manganese in the cube act as hole donors to the fourth manganese. ...
Calcium manganese oxides have captured strong scientific interest in recent years due to exceptional catalytic activities, inexpensive synthesis methods, and constitution of earth abundant and environmentally benign elements. In particular, the catalytic properties of Ca–Mn–O systems have been recognised as a potentially useful new tool in addressing energy and environmental problems. This article presents an overview of new developments in the research on Ca–Mn–O catalyst materials. The article begins with the discussion about the CaMn4O5(H2O)4 cluster which is situated within Photosystem II proteins in plant cells and is responsible for the natural water-splitting reaction. It then focuses on the crystal structures and synthesis routes of various Ca–Mn–O systems, followed by a critical discussion on the applications in catalytic water oxidation. Other known applications of Ca–Mn–O systems in the energy and environmental sectors are also discussed.
... •+ population than P D2 •+ in may proceed via different pathways depending on the S-state transitions [17][18][19] . The nature of the protonconducting O4-water chain 20 , which is composed exclusively of water molecules, is consistent with and may explain the pH-independence of proton transfer in the S 0 to S 1 transition 16 . ...
We report redox potentials (Em) for one-electron reduction for all chlorophylls in the two electron-transfer branches of, water-oxidizing enzyme photosystem II (PSII), photosystem I (PSI), and purple bacterial photosynthetic reaction centers (PbRC). In PSI, Em values for the accessory chlorophylls were similar in both electron-transfer branches. In PbRC, the corresponding Em value was 170 mV less negative in the active L-branch (BL) than in the inactive M-branch (BM), favoring B L●– formation. This contrasted with the corresponding chlorophylls, ChlD1 and ChlD2, in PSII, where Em(ChlD1) was 120 mV more negative than Em(ChlD2), implying that to rationalize electron transfer in D1-branch, ChlD1 would need to serve as the primary electron donor. Residues that contributed to Em(ChlD1) < Em(ChlD2) simultaneously played a key role in (i) releasing protons from the substrate water molecules and (ii) contributing to the larger cationic population on the chlorophyll closest to the Mn4CaO5 cluster (PD1), favoring electron transfer from water molecules. These features seem to be the nature of PSII, which needs to possess the proton-exit pathway to use a protonated electron source—water molecules.
... Whereas PSII is mostly confined to the grana regions, PSI and ATP synthase are located in the stroma lamellae (Andersson and Anderson, 1980;Dekker and Boekema, 2005;Hankamer et al., 1997;Kaftan et al., 2002). The primary electron transfer reactions take place in the reaction center of PSII, which absorbs light, oxidizes H 2 O to O 2 via the water-oxidizing complex (WOC) (Renger, 2001;Renger and Kuhn, 2007) and extracts electrons from water. During linear electron transport, these electrons are used for the reduction of plastoquinone (PQ) to plastoquinol (PQH 2 ) and are then transferred to Cytb 6 f, which mediates the electron transport to PSI via plastocyanin (PC). ...
... Protons are released in response to changes in the oxidation state (the S n state, where the subscript represents the number of oxidation steps accumulated) of the oxygen-evolving complex, and it occurs with a typical stoichiometry of 1:0:1:2 for the S 0 →S 1 →S 2 →S 3 (→S 4 )→S 0 transitions, respectively. Candidates for the relevant proton transfer pathways (e.g., (Renger 2001;Murray and Barber 2007;Ho and Styring 2008;Vassiliev et al. 2012;Ogata et al. 2013;Linke and Ho 2014) and proton-releasing sites (e.g., (Saito et al. 2015)) have been reviewed recently. The energetically lowest process for proton release from the Mn 4 CaO 5 cluster is the S 0 -to-S 1 transition, where the electron transfer occurs prior to H + release and is thus rate limiting (Dau and Haumann 2008). ...
In the cyanobacterial photosystem II (PSII), the O4-water chain in the D1 and CP43 proteins, a chain of water molecules that are directly H-bonded to O4 of the Mn4Ca cluster, is linked with a channel that connects the protein bulk surface along with a membrane-extrinsic protein subunit, PsbU (O4-PsbU channel). The cyanobacterial PSII structure also shows that the O1 site of the Mn4Ca cluster has a chain of H-bonded water molecules, which is linked with the channel that proceeds toward the bulk surface via PsbU and PsbV (O1-PsbU/V channel). Membrane-extrinsic protein subunits PsbU and PsbV in cyanobacterial PSII are replaced with PsbP and PsbQ in plant PSII. However, these four proteins have no structural similarity. It remains unknown whether the corresponding channels also exist in plant PSII, because water molecules are not identified in the plant PSII cryo-electron microscopy (cryo-EM) structure. Using the cyanobacterial and plant PSII structures, we analyzed the channels that proceed from the Mn4Ca cluster. The cyanobacterial O4-PsbU and O1-PsbU/V channels were structurally conserved as the channel that proceeds along PsbP toward the protein bulk surface in the plant PSII (O4-PsbP and O1-PsbP channels, respectively). Calculated protonation states indicated that in contrast to the original geometry of the plant cryo-EM structure, protonated PsbP-Lys166 may form a salt-bridge with ionized D1-Glu329 and protonated PsbP-Lys173 may form a salt-bridge with ionized PsbQ-Asp28 near the O1-PsbP channel. The existence of these channels might explain the molecular mechanism of how PsbP can interact with the Mn4Ca cluster.
... Also the lifetimes of the reduction of P680 +· by Y z were found to depend on the redox state of the manganese cluster (Brettel et al. 1984, Eckert and. Regardless of the detailed mechanism of water splitting (for a recent review see Renger 2001), the oxidation of two molecules of water to molecular oxygen requires the extraction of four electrons under release of four protons into the lumen. To achieve this goal the oxidising redox equivalents of four successive light induced charge separation steps are stored in the manganese cluster. ...
Im Rahmen dieser Arbeit wurde eine Apparatur zur Messung von blitzinduzierten Änderungen der Fluoreszenzquantenausbeute von photosynthetisch aktiven Proben im Zeitbereich von 100 ns bis 10 s nach einem aktinischen Laserblitz entwickelt. Es wurde gezeigt, daß diese Methode geeignet ist, um Proben mit sehr unterschiedlichem Komplexitätsgrad, wie z.B. Suspensionen von Pigment-Protein-Komplexen, Zellorganellen sowie ganze Blätter, zu analysieren. Der komplexe Zeitverlauf der Fluoreszenzquantenausbeuteänderungen, die durch einen aktinischen Blitz induziert werden, ist abhängig von den Redoxzuständen des primären Elektronendonators (P680) und des sekundären Elektronenakzeptors (QA) des Photosystems II, sowie vom Anregungszustand der Carotinoide (Car) in den Antennenkomplexen. Aufgrund der unterschiedlichen Relaxationszeiten der durch den aktinischen Blitz gebildeten Komponenten P680+?, QA- und ³Car ist es möglich, deren individuelle Beiträge zur Fluoreszenzlöschung zu separieren. Das neu entwickelte Meßsystem wurde benutzt, um den Einfluß der Lipidzusammensetzung der Thylakoidmembran auf das Reaktionsmuster des Photosystems II zu untersuchen. Dazu wurden genetisch modifizierte Pflanzen der Gattung Arabidopsis thliana benutzt, die einen reduzierten Gehalt an den Galaktolipiden Monogalactosyldiacylglycerol (MGDG) oder Digalactosyldiacylglycerol (DGDG) aufweisen. Es wurde gezeigt, daß eine Reduktion des MGDG Gehaltes um 50 % im Vergleich zum Wildtyp nicht zu einer Veränderung der transienten blitzinduzierten Änderung der Fluoreszenzquantenausbeute führte. Der DGDG Gehalt in den verschiedenen untersuchten DGDG Mutanten ist um 90 % (bzw. mehr als 90 %) reduziert. Die erhaltenen Ergebnisse zeigen, daß die an diesen Mutanten gemessenen Änderungen der Fluoreszenzquantenausbeute nicht zufriedenstellend mit dem normalerweise verwendeten Model für die Fluoreszenzlöschung durch P680+?, QA und ³Car beschrieben werden können. Es wurde gezeigt, daß das weithin akzeptierte Postulat von ungefähr gleicher Löscheffektivität für P680+? und QA revidiert werden muß. Eine Modifizierung des Standardmodels in Bezug auf diesen Punkt ermöglicht eine konsistente Beschreibung der im Wildtyp und allen Mutanten gemessenen Kinetik der transienten blitzinduzierten Fluoreszenzquantenausbeuteänderungen. Die Auswertung der Ergebnisse legt die Lokalisierung der Wirkung des DGDG-Mangels an zwei unabhängigen Orten nahe: die Antennenkomplexe und die Donatorseite des Photosystems II. In Bezug auf die Antennenkomplexe zeigte die Untersuchung, daß der relative Anteil an blitzinduzierter Bildung von ³Car in den DGDG-Mangelmutanten ansteigt. Daraus wurde die Schlußfolgerung gezogen, daß der DGDG-Mangel die strukturelle Organisation der Antennenkomplexe ändert und infolgedessen die Bildung von ³Car begünstigt. Die Wirkungsweise des DGDG-Mangels auf Photosystem II ist spezifisch für die Donatorseite des Photosystems II und betrifft nur eine Subpopulation von PS II Komplexen. In diesen modifizierten PS II Komplexen ist die schnelle Reduktion von P680+? durch YZ verlangsamt, sodaß P680+? zu einem relativ hohen Anteil durch die Rekombinationsreaktion mit QA- reduziert wird.
... Gernot had a very broad and detailed knowledge of the literature related to the primary events in photosynthesis. He summarized his insights in a large number of reviews (Renger ( , 2001(Renger ( , 2012, Renger and Renger (2008) and Messinger and Renger (2008)). Of special note is a comprehensive book 'Primary Processes of Photosynthesis' that Gernot had edited (Renger 2008b). ...
Gernot Renger (October 23, 1937–January 12, 2013), one of the leading biophysicists in the field of photosynthesis research, studied and worked at the Max-Volmer-Institute (MVI) of the Technische Universität Berlin, Germany, for more than 50 years, and thus witnessed the rise and decline of photosynthesis research at this institute, which at its prime was one of the leading centers in this field. We present a tribute to Gernot Renger’s work and life in the context of the history of photosynthesis research of that period, with special focus on the MVI. Gernot will be remembered for his thought-provoking questions and his boundless enthusiasm for science.
The photosystem II (PSII)-catalyzed water oxidation is crucial for maintaining life on earth. Despite the extensive experimental and computational research that has been conducted over the past two decades, the mechanisms of O-O bond formation and oxygen release during the S3 ∼ S0 stage remain disputed. While the oxo-oxyl radical coupling mechanism in the "open-cubane" S4 state is widely proposed, recent studies have suggested that O-O bond formation may occur from either the high-spin water-unbound S4 state or the "closed-cubane" S4 state. To gauge the various mechanisms of O-O bond formation proposed recently, the comprehensive QM/MM and QM calculations have been performed. Our studies show that both the nucleophilic O-O coupling from the Mn4 site of the high-spin water-unbound S4 state and the O5-O6 or O5-OW2 coupling from the "closed-cubane" S4 state are unfavorable kinetically and thermodynamically. Instead, the QM/MM studies clearly favor the oxyl-oxo radical coupling mechanism in the "open-cubane" S4 state. Furthermore, our comparative research reveals that both the O-O bond formation and O2 release are dictated by (a) the exchange-enhanced reactivity and (b) the synergistic coordination interactions from the Mn1, Mn3, and Ca sites, which partially explains why nature has evolved the oxygen-evolving complex cluster for the water oxidation.
Among many processes occurring in oxygenic photosynthesis, the water oxidation reaction catalyzed by the Mn4Ca cluster provides various types of insights into the field of the metal coordination chemistry. The water oxidation reaction in nature is carried out by Photosystem II (PS II), a multi subunit membrane protein complex. This light-driven reaction is made possible by a spatially separated, yet temporally connected series of cofactors along the electron transfer chain of PS II over 40 Å, through the donor—the Mn4CaO5 catalytic center, the reaction center chlorophylls, to the mobile quinone electron acceptors. Such chemical architecture provides an ideal platform to investigate how to control multi-electron/proton chemistry, using the flexibility of metal redox states, in coordination with the protein and the water network. Understanding the insights of nature's design gives inspiration of how to build artificial photosynthetic devices, where the controlled accumulation of charge and high-selectivity of products is currently challenging. The electronic and geometric structure of this catalyst have been extensively investigated, but its step-wise water oxidation mechanism is not yet completely understood. In this chapter, we summarize our current understanding of the water oxidation reaction in nature.
Energy shortage and environmental pollution limit the sustainable development of human beings. Water, as a clean and renewable resource, provides a solution for sustainable energy conversion from water oxidation catalysis. Lessons should be learned from nature to explore efficient artificial catalysts. In this chapter, we will review recent major progress in natural photosynthesis, from the structure and functions to the mechanisms for the water-oxidizing center of the biological enzyme. Later, the development of molecular and material water oxidation catalysts is discussed, with a focus on the mechanisms and rational catalyst design.
We provide a perspective on how single-molecule magnets can offer a platform to combine quantum transport and paramagnetic spectroscopy, so as to deliver time-resolved electron paramagnetic resonance at the single-molecule level. To this aim, we first review the main principles and recent developments of molecular spintronics, together with the possibilities and limitations offered by current approaches, where interactions between leads and single-molecule magnets are important. We then review progress on the electron quantum coherence on devices based on molecular magnets, and the pulse sequences and techniques necessary for their characterization, which might find implementation at the single-molecule level. Finally, we highlight how some of the concepts can also be implemented by including all elements into a single molecule and we propose an analogy between donor–acceptor triads, where a spin center is sandwiched between a donor and an acceptor, and quantum transport systems. We eventually discuss the possibility of probing spin coherence during or immediately after the passage of an electron transfer, based on examples of transient electron paramagnetic resonance spectroscopy on molecular materials.
The crucial O-O bond forming step in the water oxidising complex (WOC) of photosystem II is modelled using density functional theory calculations and compared with structural X-ray free electron laser (XFEL) determinations for the penultimate S3 state. Concerted electron flow between the Mn4O5 and Mn1O6 bonds of the complex and the nascent O-O bond are monitored using intrinsic bond orbital analysis along the reaction path. Concerted transfer to Mn1 and Mn4 of two electrons from the reactant oxos, O5 and O6, resulting in an unoccupied antibonding σ2p* orbital is the key to low barrier O-O bond formation. The potential energy surface for O-O bond formation shows a rather broad energy minimum for the oxo-oxo form ranging from 2.4 - 2.0 Å which may explain the relatively short O5-O6 bond distance reported in experimental structure studies. Alternatively the short O5-O6 bond distance may reflect a dynamic equilibrium model across the whole O-O potential energy surface.
Molecular quantum mechanics (MQM) investigations have been performed for elucidation of fundamental principles of the photo-induced water oxidation in oxygen evolving complex (OEC) of photosystem II (PSII). To this end, as a first theoretical step, broken symmetry (BS) quantum mechanics (QM) and QM(BS)/molecular mechanics (MM) calculations have been conducted for elucidation of geometrical, electronic and spin structures of the CaMn4Ox (X = 5, 6) cluster in the five steps Si (i = 0∼4) of the Kok cycle for water oxidation. The QM and QM/MM calculations have provided full optimised geometries of short-lived key intermediates and transition state structure for the O-O bond formation in the native solar-energy conversion. The interplay between theory and experiment have clearly indicated that the CaMn4O5 cluster in OEC of PSII exhibits typical physicochemical properties of strong correlation electron system (SCES) confined with effective protein field. Our QM and QM/MM computational results for key intermediates and transition structure for the O-O bond formation in the Kok cycle are now plentiful for derivation of fundamental principles (FP) for understanding of photo-induced water oxidation in OEC of PSII.
We summarize twenty nine fundamental principles (FPs) in systematic manner by QM and QM/MM calculations for understanding of water oxidation in Oxygen Evolving Complex(OEC) of Photosystem II(PSII). One of our final aims is theoretical design of the next-generation artificial photosystem materials composed of abundant metal ions.
Excitation energy transfer (EET) processes in different photosynthetic pigment-protein-complexes were analysed by time- and wavelength correlated single photon counting (TWCSPC). A new mobile 16-channel photomultiplier with flexible fiber optics, exchangeable light sources and temperature regulator (10 K – 350 K) was built up for the spectroscopy of samples in cuvettes, on surfaces or of whole leaves in vivo. The system represents a mobile setup of the powerful TCSPC technique with high optical throughput up to 106 counts/sec.
The theoretical description of the excited state dynamics in systems with pigment-pigment and pigment-protein interaction was performed by using rate equations that were applied on structures with increasing hierarchical complexity. The study started with a system consisting of two excitonically coupled Chl molecules in a tetrameric protein environment represented by the recombinant water soluble Chl binding protein (WSCP) of type IIa and it was completed with a study of the photosystem II (PSII) dynamics in whole leaves of the higher plant Arabidopsis thaliana. In this way a quantification of dissipative excited state relaxation processes as a function of increasing excitation light intensity was achieved. For parameter adaption in the corresponding systems of linear differential
equations a highly efficient algorithm was developed that allows the variation of parameters used to fit the time resolved optical data under any constraints on the coefficient matrix (e.g. invariance of thesymmetry). The approach permits the determination of selected parameter values, their probability and stability in any dynamical system. A way to calculate thermodynamic quantities (e.g. entropy) under
nonequilibrium conditions from rate equations is proposed.
The excited state dynamics observed in WSCP were explained by assuming an excitonically coupled Chl dimer that is modulated by the protein environment on different time scales. The dominating fluorescence decay component increases from 4.8 ns or 5.2 ns for Chl b or Chl a homodimers, respectively, at room temperature, to 7.0 or 6.2 ns, respectively, at 10 K. This temperature dependency is most probably caused by the pigment-pigment- interaction and the energies of the triplet states of the Chl molecules. A modulation of the electronic states of the coupled Chl dimer by the protein environment with a typical time constant of 100 ps at 10 K is inferred to be responsible for a fast and
strongly temperature dependent fluorescence component. This idea is qualitatively in line with refined theoretical models and results of complementary studies of hole burning and fluorescence line narrowing spectroscopy.
In the phycobiliprotein (PBP) antenna of the cyanobacterium Acaryochloris marina EET occurs with characteristic time constants of 400 fs, 3-5 ps and 14 ps inside trimeric phycocyanin (PC), from PC to allophycocyanin (APC) and from APC to the terminal emitter (TE) of the PBP antenna, respectively.
The TWCSPC spectra of whole cells and preparations of isolated PBP complexes exhibit a 20 ps component each that indicates the intact EET from PC to the TE in agreement with the results of transient fs absorption spectroscopy. The EET from the PBP antenna to the Chl d containing core antenna complexes of PS II represents an additional limiting transfer step of about 30-40 ps which leads to a time constant of the EET from PBP to Chl d in the range of 70 ps.
Coupled and temperature switch-able hybrid systems of surface treated CdSe/ZnS quantum dots (QDs) with 530 nm emission wavelength and the isolated PBP antenna complexes from A.marina were formed in aqueous solution by electrostatic self assembly. Based on the theory of Förster Resonance Energy Transfer (FRET) an average value of 3.2-3.5 nm was obtained for the distance between the neighbouring transition dipole moments in the QDs and the PBP antenna. It was shown that the functional coupling between QDs and PBP complexes is interrupted at temperatures below 0°C. This
effect enables the construction of switch-able molecular sensors, photosensibilisators or light harvesting devices with various applications in biochemistry, biomedicine and photovoltaics.
The approach for the analysis of some large-scale biological systems, on the base of quasiequilibrium stages is proposed. The approach allows us to reduce the detailed large-scaled models and obtain the simplified model with an analytical solution. This makes it possible to reproduce the experimental curves with a good accuracy. This approach has been applied to a detailed model of the primary processes of photosynthesis in the reaction center of photosystem II. The resulting simplified model of photosystem II describes the experimental fluorescence induction curves for higher and lower plants, obtained under different light intensities. Derived relationships between variables and parameters of detailed and simplified models, allow us to use parameters of simplified model to describe the dynamics of various states of photosystem II detailed model.
In this series of articles, the board members of ChemSusChem discuss recent research articles that they consider of exceptional quality and importance for sustainability. This entry features Prof. L. Sun, who proposes a special mechanism for O−O bond formation in photosystem II with involvement of an MnVII–oxo species induced by charge‐ and structural rearrangements. In this viewpoint, Proton transfer is involved in changes of the first coordination spheres around the MnVII–oxo site on the dangling Mn4 with de‐ and re‐coordination of carboxylates (Glu333 and Asp170).
This short communication focuses on energetics, kinetics and mechanism of oxidative photosynthetic water splitting into molecular dioxyen and four protons. The process which takes place in the Photosystem II complex of all oxygen evolving organisms comprises three different types of reaction: I) light induced generation of the strongly oxidizing cation radical P680+· ii) coupled proton and electron transfer leading to P680+· reduction by YZ, and iii) sequence of redox steps within the WOC driven by YZox. © Zhejiang University Press, Hangzhou and Springer-Verlag Berlin Heidelberg 2013.
QM(UB3LYP)/MM(AMBER) calculations were performed for the locations of the transition structure (TS) of the oxygen–oxygen (O–O) bond formation in the S4 state of the oxygen-evolving complex (OEC) of photosystem II (PSII). The natural orbital (NO) analysis of the broken-symmetry (BS) solutions was also performed to elucidate the nature of the chemical bonds at TS on the basis of several chemical indices defined by the occupation numbers of NO. The computational results revealed a concerted bond switching (CBS) mechanism for the oxygen–oxygen bond formation coupled with the one-electron transfer (OET) for water oxidation in OEC of PSII. The orbital interaction between the σ-HOMO of the Mn(IV)4–O(5) bond and the π*-LUMO of the Mn(V)1=O(6) bond plays an important role for the concerted O–O bond formation for water oxidation in the CaMn4O6 cluster of OEC of PSII. One electron transfer (OET) from the π-HOMO of the Mn(V)1=O(6) bond to the σ*-LUMO of the Mn(IV)4–O(5) bond occurs for the formation of electron transfer diradical, where the generated anion radical [Mn(IV)4–O(5)]⁻• part is relaxed to the •Mn(III)4 … O(5)⁻ structure and the cation radical [O(6)=Mn(V)1]⁺ • part is relaxed to the ⁺O(6)–Mn(IV)1• structure because of the charge-spin separation for the electron-and hole-doped Mn–oxo bonds. Therefore, the local spins are responsible for the one-electron reductions of Mn(IV)4->Mn(III)4 and Mn(V)1->Mn(IV)1. On the other hand, the O(5)⁻ and O(6)⁺ sites generated undergo the O–O bond formation in the CaMn4O6 cluster. The Ca(II) ion in the cubane- skeleton of the CaMn4O6 cluster assists the above orbital interactions by the lowering of the orbital energy levels of π*-LUMO of Mn(V)1=O(6) and σ*-LUMO of Mn(IV)4–O(5), indicating an important role of its Lewis acidity. Present CBS mechanism for the O–O bond formation coupled with one electron reductions of the high-valent Mn ions is different from the conventional radical coupling (RC) and acid-base (AB) mechanisms for water oxidation in artificial and native photosynthesis systems. The proton-coupled electron transfer (PC-OET) mechanism for the O–O bond formation is also touched in relation to the CBS-OET mechanism.
The evolution of aerobic life on earth is depended on proceeding water splitting accomplished through photosynthesis in plants, algae, and cyanobacteria. Photosystem II (PSII), with a catalytic center CaMn4O5 located on the lumenal surface, is responsible for water splitting and generating molecular oxygen through a four-step photocatalytic cycle. So far, the structure of the catalytic center and its ligation environments have been studied by different methods mostly relied on various spectroscopic techniques, disclosing unknowing aspects of the PSII components. Over the last half-decade, the experimental methods have extensively been coupled with quantum mechanics/molecular mechanics (QM/MM) methods to explore diverse aspects of PSII structure and water oxidation mechanism. However, despite the progress made in the past years, distinguishing a generally accepted mechanism on the O–O bond formation is still a challenge. This substantial challenge, if resolved, would provide a widespread criterion for development of globally deployable biomimetic model systems for water splitting catalysts. Here, we highlight some latest studies performed on the structure and function of PSII, the information that tells us how to establish new artificial catalytic systems to deliver maximum performance through water splitting in research labs.
Light driven water oxidation is a fundamental reaction in the biosphere. The Mn4Ca cluster of Photosystem II cycles through five redox states termed S0-S4 after which, oxygen is evolved. Critically, the timing of O–O bond formation within the Kok cycle remains unknown. Combining recent crystallographic, spectroscopic and DFT results, we demonstrate an atomistic S3-state model with the possibility of a low barrier to O–O bond formation prior to the final oxidation step. Furthermore, the associated one electron oxidized S4-state does not provide more advantages in terms of spin alignment or the energy of O–O bond formation. We propose that a high energy peroxide isoform of the S3-state can preferentially be oxidized by Tyrzox in the course of final electron transfer leading to O2 evolution. Such a mechanism may explain the peculiar kinetic behavior as well as serve as an evolutionary adaptation which avoids release of the harmful peroxides.
A comparison between experimental and Broken Symmetry Density Functional Theory (BS-DFT) calculated hyperfine couplings for the S2 state of the oxygen evolving complex (OEC) has been performed. The effect of Ca substitution by Sr combined with the protonation state of two terminal hydroxo or water ligands, W1 and W2, on the calculated hyperfine couplings of 55Mn, 13C, 14N, 17O and 1H nuclei has been investigated. Our findings show best agreement for OEC models which contain a hydroxo group at the W2 position and a water molecule at W1. For this model the agreement between calculated and experimental data for all hyperfine couplings is excellent. Models with a hydroxo group at W1 are particularly poor models. Sr substitution has a minor influence on calculated hyperfine couplings in agreement with experimental determinations. The sensitivity of the hyperfine couplings to relatively minor changes in the OEC structure demonstrates the power of this methodology in refining the details of its steric and electronic structure which is an essential step in formulating a complete mechanism for water oxidation by the OEC.
Nanolayered Mn oxides are among important Mn-based catalysts for water oxidation. Mn(II), (III) and (IV) ions are present in the structure, and thus, electrochemistry of the solid is very complicated. Herein, the cyclic voltammetries of the nanolayered Mn oxides in the presence of LiClO4 at pH= 6.3, under different conditions, were studied using scanning electron microscopy, transmission electron microscopy, electrochemical impedance spectroscopy, X-ray diffraction and visible spectroelectrochemistry. The scan rates, calcination temperatures and the range of the cyclic voltammetries have very important effects on the electrochemistry of nanolayered Mn oxides. The effect of D2O instead of H2O on the electrochemistry of nanolayered Mn oxide was also considered. Such nanolayered Mn oxides were reported as water-oxidizing catalysts in the presence of cerium(IV) ammonium nitrate, in the next step we studied the cyclic voltammetry of the nanolayered Mn oxide under acidic conditions and in the presence of cerium(IV) ammonium nitrate.
The cyanobacterial photosystem II (PSII) crystal structure includes more than 1300 water molecules in each monomer unit; however, their precise roles in water oxidation are unclear. To understand the origins of water molecules in the PSII crystal structure, the accessibility of bulk water molecules to channel inner spaces in PSII was investigated using the water-removed PSII structure and molecular dynamics (MD) simulations. The inner space of the channel that proceeds toward the D1-Glu65/D2-Glu312 pair (E65/E312 channel) was entirely filled with water molecules from the bulk region. In the same channel, a diamond-shaped cluster of water molecules formed near redox-active TyrZ in MD simulations. Reorientation of the D2-Leu352 side-chain resulted in formation of a hexagonal water network at the Cl-2 binding site. Water molecules could not enter the main region of the O4-water chain, which proceeds from the O4 site of the Mn4CaO5 cluster. However, in the O4-water chain, the two water-binding sites that are most distant from the protein bulk surface were occupied by water molecules that approached along the E65/E312 channel, one of which formed an H-bond with the O4 site. These findings provide key insights into the significance of the channel ends, which may utilize water molecules during the PSII photo-cycle.
This chapter describes the principles of solar energy exploitation by water splitting. The most delicate part of the overall process is the oxidation of two water molecules leading to O2 formation and H+ release. The chapter focuses on problems of this reaction and how it is performed in the multimeric photosystem II (PSII) core complex of the photosynthetic apparatus. It shows that the living organisms can use the enormous potential of highly specialized biopolymers (i.e., proteins) in tuning energetics and kinetics of the cofactors as the functional sites of biological catalysis. Of special relevance for oxidative water splitting, taking place at a Mn4OxCa cluster, is an optimized coupling between electron and proton transfer steps. The chapter considers the possibility of developing functionalized synthetic polymer matrices for both binding the catalytic sites and simultaneously providing a spatial separation of oxidative and reductive pathways of light-induced water splitting.
Sunlight is absorbed and converted to chemical energy by photosynthetic organisms. At the heart of this process is the most fundamental reaction on Earth, the light-driven splitting of water into its elemental constituents. In this way molecular oxygen is released, maintaining an aerobic atmosphere and creating the ozone layer. The hydrogen that is released is used to convert carbon dioxide into the organic molecules that constitute life and were the origin of fossil fuels. Oxidation of these organic molecules, either by respiration or combustion, leads to the recombination of the stored hydrogen with oxygen, releasing energy and reforming water. This water splitting is achieved by the enzyme photosystem II (PSII). Its appearance at least 3 billion years ago, and linkage through an electron transfer chain to photosystem I, directly led to the emergence of eukaryotic and multicellular organisms. Before this, biological organisms had been dependent on hydrogen/electron donors, such as H2S, NH3, organic acids and Fe2+, that were in limited supply compared with the oceans of liquid water. However, it is likely that water was also used as a hydrogen source before the emergence of PSII, as found today in anaerobic prokaryotic organisms that use carbon monoxide as an energy source to split water. The enzyme that catalyses this reaction is carbon monoxide dehydrogenase (CODH). Similarities between PSII and the iron- and nickel-containing form of this enzyme (Fe-Ni CODH) suggest a possible mechanism for the photosynthetic O–O bond formation.
Fluorescence (Fl) measurements provide the most popular method of estimating photosynthetic activity. [The title of Volume 19 (2004) of the Springer Series Photosynthesis and Respiration is Chlorophyll a Fluorescence: A Signature of Photosynthesis]. The main sources of fluorescence are excited Chl molecules of PSII, but fluorescence intensity is determined by a network of all the processes occurring in the photosynthetic membrane. Analyzing the time kinetics of fluorescence induction under various conditions, we can obtain information about processes occurring at different stages of energy transduction in the photosynthetic membrane (Krause and Weiss 1991; Papageorgiou et al. 2007).
The model developed here describes photosynthetic processes according to the classical Z-scheme, where PSII and PSI operate in turn (Hall and Rao 1999; Nelson and Yocum 2006). The model was developed on the basis of the simulation of electron transport processes in isolated fragments of PSI, PSII, and bacterial reaction centers (Riznichenko et al. 1990; Bukhov et al. 1988; Chrabrova et al. 1989). The distinctive feature of the model (similar to the simplified model in Chap. 11) is that it simulates the processes inside the photoreaction center complexes in more detail, taking into account also the role of electric and electrochemical potentials.
The choreography of electron transfer (ET) and proton transfer (PT) in the S-state cycle at the manganese-calcium (Mn4Ca) complex of photosystem II (PSII) is pivotal for the mechanism of photosynthetic water oxidation. Time-resolved room-temperature X-ray absorption spectroscopy (XAS) at the Mn K-edge was employed to determine the kinetic isotope effect (KIE = tauD2O/tauH2O) of the four S-transitions in a PSII membrane particle preparation in H2O and D2O buffers. We found a small KIE (1.2-1.4) for manganese oxidation by ET from Mn4Ca to the tyrosine radical (YZdot+) in the S0n->S1+ and S1n->S2+ transitions and for manganese reduction by ET from substrate water to manganese ions in the O2-evolving S3n->S0n step, but a larger KIE (~1.8) for manganese oxidation in S2n->S3+ (subscript, number of accumulated oxidizing equivalents; superscript, charge of Mn4Ca). Kinetic lag phases detected in the XAS transients prior to the respective ET steps were assigned to S3+->S3n (~150 µs, H2O; ~380 µs, D2O) and S2+->S2n (~25 µs, H2O; ~120 µs, D2O) and attributed to PT events according to their comparatively large KIE (~2.4, ~4.5). Our results suggest that proton movements and molecular rearrangements within the hydrogen-bonded network involving Mn4Ca and its bound (substrate) water ligands and the surrounding amino-acid/ water matrix govern to different extents the rates of all ET steps, but affect particularly strongly the S2n->S3+ transition, assigned as proton-coupled electron transfer (PCET). Observation of a lag phase in the classical S2->S3 transition verifies that the associated PT is a prerequisite for subsequent ET, which completes Mn4Ca oxidation to the all-Mn(IV) level.
This historical minireview describes basic lines of progress in our understanding of the functional pattern of photosynthetic water oxidation and the structure of the Photosystem II core complex. After a short introduction into the state of the art about 35 years ago, results are reviewed that led to identification of the essential cofactors of this process and the kinetics of their reactions. Special emphasis is paid on the flash induced oxygen measurements performed by Pierre Joliot (in Paris, France) and Bessel Kok (Baltimore, MD) and their coworkers that led to the scheme, known as the Kok-cycle. These findings not only unraveled the reaction pattern of oxidation steps leading from water to molecular oxygen but also provided the essential fingerprint as prerequisite for studying individual redox reactions. Starting with the S. Singer and G. Nicolson model of membrane organization, attempts were made to gain information on the structure of the Photsystem II complex that eventually led to the current stage of knowledge based on the recently published X-ray crystal structure of 3.8 Å resolution in Berlin (Germany). With respect to the mechanism of water oxidation, the impact of Gerald T. Babcock’s hydrogen abstractor model and all the considerations of electron/proton transfer coupling are outlined. According to my own model cosiderations, the protein matrix is not only a ‘cofactor holder’ but actively participates by fine tuning via hydrogen bond networks, playing most likely an essential role in water substrate coordination and in oxygen-oxygen bond formation as the key step of the overall process.
This short communication focuses on mechanistic considerations of photosynthetic water splitting, in particular on two problems: (i) coupling of proton and electron transfer of P680+• reduction by YZ, and (ii) nature of the redox steps within the WOC with special emphasis on the O-O bond formation.
Recent high-resolution crystal structures of the water-oxidizing enzyme Photosystem II (PSII) show that O4 of the catalytic Mn4CaO5 cluster forms an H-bond with a water molecule W539, which belongs to a chain of water molecules (O4-water chain). Oxidation of Mn4CaO5 to S1 resulted in elongation of the O-H bonds and decrease in pKa(O-H/O(-)) in the [O4-H…OW539-H…OW538-H…OW393] region along the O4-water chain. In S1, removal of all water molecules from the O4-water chain, except W539, resulted in a significant pKa upshift at O4: this suggests that the proton-conducting water chain serves as a conducting media for protons, and significantly decreases the donor pKa, leading to a downhill proton transfer. The absence of a corresponding proton conducting channel is disadvantageous for release of protons from the proton-releasing site, as in the case of O5 that has no H-bond partner.
To mimic the electron donor side of Photosystem II (PSII), a number of supramolecular model complexes have been designed and synthesized. Ruthenium(II) tris-bipyridyl complexes have been used in most cases as photosensitizers, mimicking the function of P-680 in PSII. As electron donors, monomeric and dimeric manganese complexes and tyrosine have been introduced into the supramolecular systems, modelling the Mn cluster and Tyrosinez respectively in PSII For monomeric manganese complexes, di-, tri- and tetradentate ligands have been linked to a Ruthenium(II) trisbipyridyl type complex; and for dimeric manganese complexes hepta-dentate ligands containing pyridines have been used. Some related ligands, where two pyridines have been replaced by phenolate groups have also been synthesized, in order to get ligands that can stabilize manganese complex in high valence states. Photophysical and photochemical studies showed that the electron transfer rate from monomeric Mn complex to photo-generated Ru(III) was low when Mn-Ru distance was long, while the electron transfer rate was enhanced when the Mn-Ru distance was short. However, the excited state of ruthenium complex was quenched if Mn got close to the Ru, leading to a short lifetime. By synthetically inserting a tyrosine unit between Ru and Mn moieties, quenching of the excited state lifetime of Ru was reduced, and the electron transfer from Mn to Ru(III) was very fast although the Mn-Ru distance was long. These supramolecular Ru-Mn systems are closely modelling the electron donor side of PSII both functionally and structurally. The design and synthesis of these model systems are summarized and discussed in this chapter.
In this chapter, we give an overview of nature's singular biological process for producing dioxygen by the oxidation of water in photosynthetic organisms. The harnessing of light to accomplish the splitting of water was arguably nature's most successful experiment in biological innovation. It enabled global proliferation of oxygenic photosynthesis and created the biogeochemical cycles of oxygen and carbon on earth. We review the atomic structure of the metalloenzyme, the photosystem II water oxidation complex (PSII-WOC), as revealed by X-ray diffraction and spectroscopic techniques. We describe (i) the electronic and geometric structure of its inorganic core, Mn4CaO5, based upon spectroscopic and diffraction measurements; (ii) the organization of the protein subunits as revealed by X-ray diffraction and the role of select amino acid residues by site-directed mutagenesis; and (iii) the kinetics of the formation of light-induced intermediates, the exchange rates between substrate and free water molecules, the stoichiometry of electron and proton release, and the activation energies to discuss possible mechanisms for photosynthetic water splitting. The chemistry of photosynthetic water splitting is energetically demanding and mechanistically complex. The lessons learned from nature have guided chemists seeking to incorporate these design principles within catalysts suitable for abiotic water splitting. We end with a description of the few synthetic manganese complexes that have been found to produce oxygen from water and discuss the chemical mechanisms by which they appear to function.
Keywords:
bioinorganic chemistry;
calcium;
catalysis;
manganese;
oxygen evolution;
photosynthesis;
photosystem II;
water splitting
In this article, we give an overview of nature's singular biological process for producing oxygen gas by the oxidation of water in photosynthetic organisms. The harnessing of light to accomplish the splitting of water was arguably nature's most successful experiment in biological innovation. It enabled global proliferation of oxygenic photosynthesis and created the biogeochemical cycles of oxygen and carbon on earth. We review the atomic structure of the metalloenzyme, the photosystem II water splitting complex (PSII-WOC), as revealed by X-ray diffraction and spectroscopic techniques. We describe: 1) The electronic structure of its inorganic core, Mn4Ca1OxCl1–2(HCO3)y, based upon spectroscopic and magnetic susceptibility measurements, 2) the organization of the protein subunits as revealed by X-ray diffraction and the role of select amino acid residues as revealed by site-directed mutagenesis, and 3) the kinetic sequence of steps during assembly of the inorganic core to the cofactor-depleted apo-protein and the functional consequences of substitution of the inorganic cofactors. We use these data together with physico-chemical data describing the formation of light-induced intermediates, the exchange rates between substrate and free water molecules, the stoichiometry of electron and proton release and the activation energies to discuss possible mechanisms for photosynthetic water splitting. The chemistry of photosynthetic water splitting is energetically demanding and mechanistically complex. The lessons learned from nature have guided chemists seeking to incorporate these design principles within catalysts suitable for abiotic water splitting. We end with a description of the few synthetic manganese complexes that have been found to produce oxygen gas from water and discuss the chemical mechanisms by which they appear to function.
Keywords:
bioinorganic chemistry;
calcium;
catalysis;
manganese;
oxygen evolution;
photosynthesis;
photosystem II;
water splitting
A new high valent complex [Mn2(III, III)L(μ-OAc) 2]·PF6(2a) was prepared, where L was the trianion of 2, 6-bis {[(2-hydroxy-5-tert-butylbenzyl) (pyridyl-2-methyl)-amino]-methyl}- 4-methylphenol, which contains two additional phenolate groups and two tert-butyl groups compared to its parent [Mn2(II, II) (bpmp) (μ-OAc)2]·ClO4 (1). These improvements narrowed the disparity between the new model and (Mn)4 cluster (OEC in nature). Moreover, L was modified to be covalently linked with Ru(II) tris-bipyridine through an amide bond to construct a complex 2b for the study of photoinduced electron transfer (PET). UV-vis, IR, emission spectra and electrochemistry were used to investigate their photochemistry properties. The results showed that 2b has good photochemistry properties and the E 1/2 of Ru3+/Ru2+ was higher than those of phenol+/phenol and Mn (III, IV)/Mn (III, III). After coordination of manganese ions, the electron transfer process in the model complex conforms to the basic principles of electron donor side of photosystem II (PS II) in nature.
The influencing factors on heterogeneous water oxidation catalysis (WOC) were investigated in a synthetic photosystem II model developed by adsorbing [(OH2)(terpy)MnIII(μ-O)2MnIV(terpy)(OH2)]3+ (terpy = 2,2′:6′,2″-terpyridine) (1) as an oxygen evolving center onto mica. For chemical WOC using a Ce4+ oxidant, the catalytic activity of 1 on mica increased by a factor of 2.3 or 1.4 by co-adsorption (0.015 mmol g−1) of redox-inactive trications of Al3+ or Ce3+ with 1 (0.15 mmol g−1), respectively, whereas it decreased by co-adsorption (0.25 mmol g−1) of excess Al3+ or Ce3+. The cooperative catalysis by two equivalents of the adsorbed 1 for water oxidation could be facilitated by enrichment of 1 by trications at their low co-adsorption conditions. The decreased catalytic activity at high trications co-adsorption conditions could be explained by impeded penetration of Ce4+ oxidant ions into a mica interlayer. For photochemical WOC containing a [Ru(bpy)3]2+ (bpy = 2,2′-bipyridine) photoexcitation center in mica, the drying treatment at 65 °C under the vacuum after 1 adsorption was required in adsorbate preparation, possibly to maintain favorable arrangement of 1 and [Ru(bpy)3]2+ in a mica interlayer. The drying treatment at 65 °C under the vacuum after [Ru(bpy)3]2+ adsorption inactivated the photochemical WOC. The proton-coupled electron transport from interior [Ru(bpy)3]2+ centers to ones near the surface in mica is considered to be suppressed by the drying treatment, which could be responsible for the inactivated photochemical WOC.
This minireview summarizes our current knowledge on structure, energetics and the functional mechanisms of the reaction centres (RCs) of non-oxygenic photosynthesis and Photosystems I and II of oxygenic photosynthesis. At the RCs, the key steps of transformation of solar radiation into electrochemical free energy take place. Two types of RCs exist which differ in the nature of their electron acceptor for 'stable' light induced charge separation: iron-sulphur cluster in type I and quinone in type II. The type II of oxygenic photosynthesis is unique since it has the added machinery for oxygen evolution.
Flash-induced redox reactions in spinach PS II core particles were investigated with absorbance difference spectroscopy in the UV-region and EPR spectroscopy. In the absence of artificial electron acceptors, electron transport was limited to a single turnover. Addition of the electron acceptors DCBQ and ferricyanide restored the characteristic period-four oscillation in the UV absorbance associated with the S-state cycle, but not the period-two oscillation indicative of the alternating appearance and disappearance of a semiquinone at the QB-site. In contrast to PS II membranes, all active centers were in state S1 after dark adaptation. The absorbance increase associated with the S-state transitions on the first two flashes, attributed to the Z(+)S1→ZS2 and Z(+)S2→ZS3 transitions, respectively, had half-times of 95 and 380 μs, similar to those reported for PS II membrane fragments. The decrease due to the Z(+)S3→ZS0 transition on the third flash had a half-time of 4.5 ms, as in salt-washed PS II membrane fragments. On the fourth flash a small, unresolved, increase of less than 3 μs was observed, which might be due to the Z(+)S0→ZS1 transition. The deactivation of the higher S-states was unusually fast and occurred within a few seconds and so was the oxidation of S0 to S1 in the dark, which had a half-time of 2-3 min. The same lifetime was found for tyrosine D(+), which appeared to be formed within milliseconds after the first flash in about 10% inactive centers and after the third and later flashes by active centers in Z(+)S3.
Redox changes of the oxygen evolving complex in PS II core particles were investigated by absorbance difference spectroscopy in the UV-region. The oscillation of the absorbance changes induced by a series of saturating flashes could not be explained by the minimal Kok model (Kok et al. 1970) consisting of a 4-step redox cycle, S0 → S1 → S2 → S3 → S0, although the values of most of the relevant parameters had been determined experimentally. Additional assumptions which allow a consistent fit of all data are a slow equilibration of the S3 state with an inactive state, perhaps related to Ca(2+)-release, and a low quantum efficiency for the first turnover after dark-adaptation. Difference spectra of the successive S-state transitions were determined. At wavelengths above 370 nm, they were very different due to the different contribution of a Chl bandshift in each spectrum. At shorter wavelengths, the S1 → S2 transition showed a difference spectrum similar to that reported by Dekker et al. 1984b and attributed to an Mn(III) to Mn(IV) oxidation. The spectrum of absorbance changes associated with the S2 → S3 transition was similar to that reported by Lavergne 1991 for PS II membranes. The S0 → S1 transition was associated with a smaller but still substantial absorbance increase in the UV. Differences with the spectra reported by Lavergne 1991 are attributed to electrostatic effects on electron transfer at the acceptor side associated with the S-state dependence of proton release in PS II membranes.
Progress in photosynthesis research is strongly dependent on instrumentation. It is therefore not surpr- ing that the impressive advances that have been made in recent decades are paralleled by equally impressive advances in sensitivity and sophistication of physical equipment and methods. This trend started already shortly after the war, in work by pioneers like Lou Duysens, the late Stacy French, Britton Chance, Horst Witt, George Feher and others, but it really gained momentum in the seventies and especially the eighties when pulsed lasers, pulsed EPR spectrometers and solid-state electronics acquired a more and more prominent role on the scene of scientific research. This book is different from most others because it focuses on the techniques rather than on the scientific questions involved. Its purpose is three-fold, and this purpose is reflected in each chapter: (i) to give the reader sufficient insight in the basic principles of a method to understand its applications (ii) to give information on the practical aspects of the method and (iii) to discuss some of the results obtained in photosynthesis research in order to provide insight in its potentalities. We hope that in this way the reader will obtain sufficient information for a critical assessment of the relevant literature, and, perhaps more important, will gain inspiration to tackle problems in his own field of research. The book is not intended to give a comprehensive review of photosynthesis, but nevertheless offers various views on the exciting developments that are going on.
The cornerstone of the evolution of highly organized forms of living matter was the “invention” of a biomolecular device in the early biosphere that was able to dissociate water into dioxygen and metabolically bound hydrogen by using visible light. This event, which occurred in photosynthesizing organisms about 2–3 billion years ago, had two consequences of paramount importance: (a) it allowed the huge water pool on the earth’s surface to become available as a hydrogen source for the biosphere, and (b) the resulting formation of dioxygen as a photosynthetic “waste” product led to the present day aerobic atmosphere and as consequence to the formation of the protective ozone layer. The advent of O2 in the atmosphere opened the road for the much more efficient exploitation of the free energy content of food through aerobic respiration of heterotrophic organisms. This immense bioenergetic advantage which can hardly be overestimated, however, had to be paid for by a markedly enhanced risk to the organism of damage due to the formation of very reactive oxygen radicals. Protective mechanisms are therefore indispensible in order to survive in an aerobic atmosphere (for further reading, see ref. 1).
Photobiology comprises all phenomena that arise from the interaction of living matter with electromagnetic radiation in the wavelength region of near ultraviolet (NUV), visible (VIS) and near infrared (NIR). This field is a very important area in life sciences because a large number of processes in biological systems are energetically driven and/or mechanistically regulated by solar radiation. Likewise, light is also of central relevance for transfer and exchange of information (bioluminescence, process of vision etc.). With respect to solar energy exploitation, photosynthesising organisms are of paramount importance and therefore a unique topic of photobiology. Apart from its role as driving force for photosynthesis, light also regulates the formation of its microscopic apparatus as well as the macroscopic shape of plants (photomorphogenesis). Further, too much light can exert deleterious effects via mechanisms referred to as photoinhibition and photodegradation.
The key evolutionary step in the exploitation of solar radiation by photosynthetic water cleavage was the “invention” of a molecular device that enables the light-driven oxidation of two water molecules to molecular oxygen. Two indispensable prerequisites are required to perform water oxidation: (a) the generation of sufficiently oxidising redox equivalents and (b) the cooperation of four oxidising redox equivalents. This goal was achieved about 2-3 billion years ago at the level of prokaryotic photosynthesising organisms. The result was a multimeric pigment protein complex referred to as Photosystem II (PS II) that is anisotropically incorporated in the thylakoid membrane and acts as a water-plastoquinone oxidoreductase (for a recent review see ref. [1]). The overall reaction of PS II leading to formation of membrane bound plastoquinol (PQH2) and release of molecular oxygen into the atmosphere comprises three reaction sequences: i) photooxidation of a special Chla component (symbolised by P680) and subsequent stabilisation of the primary charge separation by rapid electron transfer from Pheo-· to a specially bound plastoquinone-9 molecule (QA) (for a review see ref. [2]); ii) cooperation of four strongly oxidising holes via a sequence of four univalent redox steps at a manganese containing unit, the water oxidising complex (WOC) that leads to molecular oxygen and four protons (for reviews see refs. [3-5]), and iii) cooperation of two reducing equivalents (electrons) via a sequence of two univalent redox steps at a plastoquinone-9 molecule (QB), transiently bound into a protein pocket (QB-site). This reaction leads to PQH2 formation (for reviews see refs. [6, 7]).
Water cleavage into dioxygen and four protons requires the cooperation of four redox equivalents of sufficient oxidizing power. In photosynthesis the 1-electron oxidant is generated by electron ejection from the excited singlet state of a special chlorophyll a complex (P680) with pheophytin a (Pheo) as primary acceptor and the indispensible stabilization through Pheo− reoxidation by a specifically bound plastoquinone (QA). The oxidizing equivalents are transferred from P680+ via a tyrosine residue (Yz) into a manganese containing hole storage unit referred to as HSU(Mn). After accumulation of four holes by a sequential univalent storage process (described by S0→S1→S2→S3→S4) oxygen is evolved and HSU(Mn) returns to state S0 (for review see ref. 1,2).
The influence of H/D-exchange on the electron transfer from YZ to P680+•, and the recombination reaction between P680+• and QA-• in Tris-treated photosystem 2 (PS2) membrane fragments at pL [L = lyonium ion (H,D)] = 6.5 was investigated by monitoring and numerical analysis of flash-induced absorption changes at 830 and 320 nm, respectively. The H/D-exchange caused retardation by a factor of approximately 3 of the electron transfer from YZ to P680+•. In marked contrast, no significant effect was observed on the kinetics of P680+•QA-•* charge recombination. In addition, the pH-dependence of P680+•/QA-• recombination kinetics were analysed in samples where YZ was functionally eliminated by exposure of Tris-treated PS2 fragments to strong irradiance. In this case the relaxation kinetics could be fitted by three-exponentials with half lifetimes of 150 μs (fast), 800 μs (middle) and 10 ms (slow) at pH = 6.0. The fast and middle kinetics were only slightly dependent on pH in the range from 5.0 to 8.0. On the other hand, the normalised amplitudes of these kinetics were markedly pH-dependent. Furthermore, the normalised extent of the slow kinetics was significantly larger in the absorption changes at 320 nm, reflecting the turnover of QA, than at 830 nm as an indicator of P680+• formation and decay. One possible explanation of this feature is provided by an assumption that Tris-washed PS2 membrane fragments exposed to a strong irradiance contain a redox component competing with QA-• in the reduction of P680+•. Furthermore, the pH-dependent changes of the overall kinetics of P680+•QA-• recombination originated predominantly from different ratios of the extent of the fast and middle components rather than from marked modifications of the rate constants.
The possibility to determine the difference spectra Δεi+1jλ of each univalent redox step Si→Si+1(i=0,...3) of the water-oxidizing enzyme system was analyzed by theoretical calculations and by measurements of 320 nm absorption changes induced by a train of saturating laser flashes (FWHM:7 ns) in PS II membrane fragments. It was found: a) Lipophilic quinones complicate the experimental determination of optical changes due the Si-state transitions because they lead to an additional binary oscillation probably caused by a reductant-induced oxidation of the Fe2+ at the PS II acceptor side. b) In principle, a proper separation can be achieved at sufficiently high K3[Fe(CN)6] concentrations. c) An unequivocal deconvolution into the difference spectra Δεi+1jλ of flash train-induced optical changes which are exclusively due to Si-state transitions is impossible unless the Kok parameters α, β and [Si]0 can be determined by an independent method.
Measurements of the oxygen yield induced by a flash train reveals, that in thylakoids and PS II membrane fragments Si is the stable state of dark adapted samples even at alkaline pH (up to pH=9). However, in PS II membrane fragments at pH>7.7 the misses probability α markedly increases, in contrast to the properties of intact thylakoids. Based on these data the possibility is discussed that an equilibrium exists of two types of S2-states with different properties.
The functional properties of a purified homogeneous spinach PS II-core complex with high oxygen evolution capacity (Haag et al. 1990a) were investigated in detail by measuring thermoluminescence and oscillation patterns of flash induced oxygen evolution and fluorescence quantum yield changes. The following results were obtained:a)
Depending on the illumination conditions the PS II-core complexes exhibit several thermoluminescence bands corresponding to the A band, Q band and Zv band in PS II membrane fragments. The lifetime of the Q band (Tmax=10°C) was determined to be 8s at T=10°C. No B band corresponding to S2QB− or S3QB− recombination could be detected.
b)
The flash induced transient fluorescence quantum yield changes exhibit a multiphasi relaxation kinetics shich reflect the reoxidation of Q
A−. In control samples without exogenous acceptors this process is markedly slower than in PS II membrane fragments. The reaction becomes significantly retarded by addition of 10 μM DCMU. After dark incubation in the presence of K3[Fe(CN)6
c)
Excitation of dark-adapted samples with a train of short saturating flashes gives rise to a typical pattern dominated by a high O2 yield due to the third flash and a highly damped period four oscillation. The decay of redox states S2 and S3 are dominated by short life times of 4.3 s and 1.5 s, respectively, at 20°C.
The results of the present study reveal that in purified homogeneous PS II-core complexes with high oxygen evolution isolated from higher plants by β-dodecylmaltoside solubilization the thermodynamic properties and the kinetic parameters of the redox groups leading to electron transfer from water to QA are well preserved. The most obvious phenomenon is a severe modification of the QB binding site. The implications of this finding are discussed.
Old and very recent experiments on the extent and the rate of proton release during the four reaction steps of photosynthetic water oxidation are reviewed. Proton release is discussed in terms of three main sources, namely the chemical production upon electron abstraction from water, protolytic reactions of Mn-ligands (e.g. oxo-bridges), and electrostatic response of neighboring amino acids. The extent of proton release differs between the four oxidation steps and greatly varies as a function of pH both, but differently, in thylakoids and PS II-membranes. Contrastingly, it is about constant in PS II-core particles. In any preparation, and on most if not all reaction steps, a large portion of proton transfer can occur very rapidly (<20 μs) and before the oxidation of the Mn-cluster by Yz (+) is completed. By these electrostatically driven reactions the catalytic center accumulates bases. An additional slow phase is observed during the oxygen evolving step, S3⇒S4→S0. Depending on pH, this phase consists of a release or an uptake of protons which accounts for the balance between the number of preformed bases and the four chemically produced protons. These data are compatible with the hypothesis of concerted electron/proton-transfer to overcome the kinetic and energetic constraints of water oxidation.
Solar energy exploitation by photosynthetic water cleavage is of central relevance for the development and sustenance of all higher forms of living matter in the biosphere. The key steps of this process take place within an integral protein complex referred to as Photosystem II (PS II) which is anisotropically incorporated into the thylakoid membrane. This minireview concentrates on mechanistic questions related to i) the generation of strongly oxidizing equivalents (holes) at a special chlorophyll a complex (designated as P680) and ii) the cooperative reaction of four holes with two water molecules at a manganese containing unit WOC (water oxidizing complex) resulting in the release of molecular oxygen and four protons. The classical work of Pierre Joliot and Bessel Kok and their coworkers revealed that water oxidation occurs via a sequence of univalent oxidation steps including intermediary redox states Si (i = number of accumulated holes within the WOC). Based on our current stage of knowledge, an attempt is made a) to identify the nature of the redox states Si, b) to describe the structural arrangement of the (four) manganese centers and their presumed coordination and ligation within the protein matrix, and c) to propose a mechanism of photosynthetic water oxidation with special emphasis on the key step, i.e. oxygen-oxygen bond formation. It is assumed that there exists a dynamic equilibrium in S3 with one state attaining the nuclear geometry and electronic configuration of a complexed peroxide. This state is postulated to undergo direct oxidation to complexed dioxygen by univalent electron abstraction with YZox and simultaneous internal ligand to metal charge transfer.
Key questions on the mechanism will be raised. The still fragmentary answers to these questions not only reflect our limited knowledge but also illustrate the challenges for future research.
A comparative study of X-band EPR and ENDOR of the S2 state of photosystem II membrane fragments and core complexes in the frozen state is presented. The S2 state was generated either by continuous illumination at T=200 K or by a single turn-over light flash at T=273 K yielding entirely the same S2 state EPR signals at 10 K. In membrane fragments and core complex preparations both the multiline and the g=4.1 signals were detected with comparable relative intensity. The absence of the 17 and 23 kDa proteins in the core complex preparation has no effect on the appearance of the EPR signals. (1)H-ENDOR experiments performed at two different field positions of the S2 state multiline signal of core complexes permitted the resolution of four hyperfine (hf) splittings. The hf coupling constants obtained are 4.0, 2.3, 1.1 and 0.6 MHz, in good agreement with results that were previously reported (Tang et al. (1993) J Am Chem Soc 115: 2382-2389). The intensities of all four line pairs belonging to these hf couplings are diminished in D2O. A novel model is presented and on the basis of the two largest hfc's distances between the manganese ions and the exchangeable protons are deduced. The interpretation of the ENDOR data indicates that these hf couplings might arise from water which is directly ligated to the manganese of the water oxidizing complex in redox state S2.
The functional connection between redox component Yz
identified as Tyr-161 of polypeptide D-1 (Debus et al. 1988) and P680+ was analyzed by measurements of laser flash induced absorption changes at 830 nm in PS II membrane fragments from spinach. It was found that neither DCMU nor the ADRY agent 2-(3-chloro-4-trifluoromethyl) anilino-3,5-dinitrothiophene (ANT 2p) affects the rate of P680+ reduction by Yz
under conditions where the catalytic site of water oxidation stays in the redox state S1. In contrast to that, a drastic retardation is observed after mild trypsin treatment at pH=6.0. This effect which is stimualted by flash illumination can be largely reversed by Ca2+. The above mentioned data lead to the following conclusions: (a) the segment of polypeptide D-1 containing Tyr-161 and coordination sites of P680 is not allosterically affected by structural changes due to DCMU binding at the QB-site which is also located in D-1. (b) ANT 2p as a strong protonophoric uncoupler and ADRY agent does not modify the reaction coordinate of P680+ reduction by Yz
, and (c) Ca2+ could play a functional role for the electronic and vibrational coupling between the redox groups Yz
and P680. The electron transport from Yz
to P680+ is discussed within the framework of a nonadiabatic process. Based on thermodynamic considerations the reorganization energy is estimated to be in the order of 0.5 V.
A number of carboxyl groups in turkey ovomucoid third domain (OMTKY3) have low pKa values. A previous study suggested that neighboring amino groups were primarily responsible for the low carboxyl pKa values. However, the expected elevation in pKa values for these amino groups was not observed. In the present study, site-directed mutagenesis is used to investigate the origins of perturbed carboxyl pKa values in OMTKY3. Electrostatic calculations suggest that Lys 34 has large effects, 0.4−0.6 unit, on Asp 7, Glu 10, and Glu 19 which are 5−11 Å away from Lys 34. Two-dimensional 1H NMR techniques were used to determine pKa values of the acidic residues in OMTKY3 mutants in which Lys 34 has been replaced with threonine and glutamine. Surprisingly, the pKa values in the mutants are very close to those of the wild-type protein. The insensitivity of the acidic residues to replacement of Lys 34 suggests that long-range electrostatic interactions play less of a role in perturbing carboxyl pKa values than originally thought. We hypothesize that hydrogen bonds play a key role in perturbing some of the carboxyl ionization equilibria in OMTKY3.
Protein-bound [FeS] clusters function widely in biological electron-transfer reactions, where their midpoint potentials control both the kinetics and thermodynamics of these reactions. The polarity of the protein environment around [FeS] clusters appears to contribute largely to modulating their midpoint potentials, with local protein dipoles and water dipoles largely defining the polarity. The function of the [4Fe-4S] cluster containing Fe protein in nitrogenase catalysis is, at least in part, to serve as the nucleotide-dependent electron donor to the MoFe protein which contains the sites for substrate binding and reduction. The ability of the Fe protein to function in this manner is dependent on its ability to adopt the appropriate conformation for productive interaction with the MoFe protein and on its ability to change redox potentials to provide the driving force required for electron transfer. Phenylalanine at position 135 is located near the [4Fe-4S] cluster of nitrogenase Fe protein and has been suggested by amino acid substitution studies to participate in defining both the midpoint potential and the nucleotide-induced changes in the [4Fe-4S] cluster. In the present study, the crystal structure of the Azotobacter vinelandii nitrogenase Fe protein variant having phenylalanine at position 135 substituted by tryptophan has been determined by X-ray diffraction methods and refined to 2.4 Angstrom resolution. A comparison of available Fe protein structures not only provides a structural basis for the more positive midpoint potential observed in the tryptophan substituted variant but also suggests a possible general mechanism by which the midpoint potential could be controlled by nucleotide interactions and nitrogenase complex formation.
The decay kinetics for the S2 and S3 states of the oxygen-evolving complex in Photosystem II have been measured in the presence of an external electron acceptor. The S2- and S3-states decay monophasically with half-decay times at 18°C of 3–3.5 min and 3.5–4 min, respectively. The results also show that S3 decays via S2 under these circumstances. The temperature dependence of the individual S-state transitions has been measured in single flash experiments in which the multiline EPR signal originating from the S2 state has been used as spectroscopic probe. The half-inhibition temperatures are for S0 to S1 220–225 K, for S1 to S2 135–140 K, for S2 to S3 230 K and for the S3-to-S0 transition 235 K.
The temperature dependence of S-state transition kinetics was measured by spectroscopy in the ultraviolet region with O2-evolving Photosystem II particles from a thermophilic cyanobacterium. By proper selection of ferricyanide concentration and the measuring wavelength, the absorption changes due to S-state transitions were explicitly measured with minimized superposition of binary absorption changes due to acceptor side reactions. The half-times of S1 → S2, S2 → S3 and S3 → S0 transitions were 60, 60 and 800 μs, respectively, at 50°C, the optimal temperature for oxygen evolution by the particles, but were slowed down to 70, 120–150 and 1300 μs at 25°C, and 106, 300 and 5500 μs at 1°C, respectively. In the whole range of 1–50°C the Arrhenius plots showed no break or discontinuity for S1 → S2 and S2 → S3 transitions, with apparent activation energies of 9.6 and 26.8 kJ/mol, respectively. The Arrhenius plot for S3 → S0 transition, however, was composed of two straight lines with a clear break at 16°C, and the apparent activation energies above and below the break temperature were 15.5 and 59.4 kJ/mol, respectively. The implications of these data and especially of the break temperature for the S3 → S0 transition were discussed.
The effect of mild trypsination on the system-II reaction pattern of inside-out thylakoids has been analyzed by measurements of oxygen yield, fluorescence induction and laser-pulse-induced absorption changes at 320 and 830 nm. The following was found. (1) The average oxygen yield per flash drastically declines after trypsination at pH 7.4, while at pH 6.5 only small effects are observed. (2) The area over the fluorescence induction curves becomes reduced by 30–40% after trypsination at either pH 6.5 or 7.4, but in the latter case the maximum level is attained only after addition of hydroxylamine as PS-II donor. On the other hand, the area over the induction curve in the presence of DCMU, which is 10–12 times smaller than without DCMU, remains unaffected by trypsin treatment. (3) The oscillation pattern of the oxygen yield induced by a flash train in dark-adapted inside-out thylakoids is not markedly affected by trypsin treatment, even at more than 80% inhibition of the oxygen-evolving capacity. (4) After trypsination of inside-out thylakoids, a large 10 μs decay arises in the relaxation kinetics of the 830 nm absorption changes, whereas the 320 nm absorption changes are dominated by a rather slow decay. (5) The half-life time of the microsecond kinetics at 830 nm elicited by trypsination of inside-out thylakoids reveals almost the same pH dependence as the corresponding relaxation kinetics in Tris-washed inside-out thylakoids. (6) The relaxation kinetics of the absorption changes at 320 and 830 nm in Tris-washed inside-out thylakoids become significantly modified after trypsin treatment. Based upon these findings, it is concluded that beyond the well-characterized polypeptides with 16, 23 and 33 kDa there exists a further protein that is exposed to the inner side of the thylakoid and that affects the electron transport in system II. The nature and the physiological role of this polypeptide still remain to be elucidated.
The protonization pattern of the endogenous donor component D1 which feeds electrons directly into chl-a+II has been analyzed in Tris-washed inside-out thylakoids with the aid of appropriate pH-indicators. It was found that under repetitive flash excitation the amount of protons released is proportional to the extent of D1-oxidation, depending on the time between the flashes. The kinetics of D1-oxidation (being practically the same as in normal Tris-washed chloroplasts) are faster than the proton release by two orders of magnitude. The results lead to the conclusion that D1 is protonized in the reduced state with pK(Dox1) < 5 and becomes deprotonized in the oxidized state with pK(Dred1) ≳ 8. The proton release is kinetically limited by a transport barrier. Implications on the interpretation of the proton release pattern in preparation with intact water oxidation are discussed.
In Tris-washed Photosystem-II particles we are able to induce an EPR signal in the dark by addition of an iridium salt (K2IrCl6). This signal is attributed to signal IIs (slow) (D+) and the redox titration gives an Em value of 760 mV for the couple . On the basis of our previous studies on the equilibrium between D+Z and DZ+ (K = 104) (Boussac, A. and Etienne, A.L. (1982) Biochem. Biophys. Res. Commun. 109, 1200–1205), we therefore attribute a value of 1 V for the Em of the couple. A second effect of K2IrCl6 is to modify the spectral characteristics of signal II. We conclude that K2IrCl6 is able to change the environment of the species from which signal IIs and signal IIf originate.
(1) The first part of this paper is devoted to methodological tests pertaining to the deconvolution of the spectra of the S-transitions from flash sequence data, (i) The spectrum and decay kinetics of 'inactive PS II centers' were analyzed, showing similarity with normal centers inhibited on the QB site. When this contribution is subtracted, the change on the first flash agrees with estimate of the S1→S2 change derived from deconvolution. (ii) Preillumina-tion procedures in PS II (BBY) particles, aimed at modulating the initial S0/S1 distribution are confirmed to express deactivation towards the S1 state, with no involvement of an ‘S−1’ state, (iii) Elimination of semiquinone binary oscillations in BBY's is achieved by allowing total reoxidation by DCBQ after each flash, (iv) A deconvolution treatment is described, using the difference between two sequences. This method allows a determination of the initial S0/S1 distributions that cross-checks the conclusions (ii). (v) Oscillations of the amplitude of the ms-phase of the 295 nm absorption changes are shown to differ significantly from those corresponding to the S-transitions. This phase is expected to reflect predominantly the O2 release reaction, with slight deviations due to other transitions. Deconvolution results are in satisfactory agreement with this prediction. (2) The upshot of this work is to present improved spectra of the S-transitions. The basic features that were previously established with different material and deconvolution method are confirmed: negligible UV change on S0→S1, and significantly different spectra for the two other transitions. A shoulder around 350 nm on the S1→S2 spectrum and a secondary peak in the same region for the S2→S3 spectrum are now resolved. The S0→S1 step causes an electrochromic shift in the blue region, with direction opposite to S1→S2. Interpretation of these results is discussed.
Electron-transfer reactions between ions and
molecules in solution have been the subject of
considerable experimental study during the past
three decades. Experimental results have also been
obtained on related phenomena, such as reactions
between ions or molecules and electrodes, charge-transfer
spectra, photoelectric emission spectra of
ionic solutions, chemiluminescent electron transfers,
electron transfer through frozen media, and
electron transfer through thin hydrocarbon-like
films on electrodes.
The Ca2+-depleted photosystem II (PS II) was studied by pulsed EPR spectroscopy. A short (5 s) illumination of the PS II in the S′2 state resulted in formation of the doublet S′3 signal at g ≈ 2.02 with a splitting of about 15 mT concomitant with the decrease of the modified Mn multiline signal by about 40%. Further illumination up to three minutes led to the formation of another S′3 signal with a singlet-like feature at g ≈ 1.98 and to the complete disappearance of the modified Mn multiline signal. Computer simulations of the shape of the doublet S′3 signal and the dependence of its electron spin echo amplitude on the microwave field strength suggest that dipole and exchange interactions between two organic radicals are responsible for the doublet S′3 signal. The pulsed ENDOR-induced EPR measurements indicate that the ENDOR spectrum of the tyrosine radical YZ+ is associated with the doublet S′3 signal only.
A theoretical method to extract structural information on spin-exchange- coupled manganese tetramers from the EPR spectroscopy data is presented. This method has been applied to two EPR data, i.e. the ground-state spin (S = 1/2) and the first excitation energy (30-37 cm-1), for the S2-state Mn tetramer in the photosynthetic oxygen-evolving complex, which exhibits a 'g = 2 multiline' EPR signal. Based on the EXAFS data, and the manganese chemistry, a simplified model spin Hamiltonian to describe the S2-state Mn cluster will be presented, such that two spin-exchange interactions due to 2.7-2.8 Å and/or 3.2-3.5 Å Mn-Mn bonds can vary from weak to strong coupling, sensitively, depending upon the bridge structure, except for a strong- antiferromagnetic interaction due to a 2.7 Å Mn(III)-Mn(IV) bond and the other weak ones. By computer-search of the possible spin-exchange structures with respect to these two parametric interactions, it was found that (1) a dimer of di-μ2-oxo bridged Mn dimers, a propeller-type tetramer in which the central Mn ion is chelated by three di-μ-oxo Mn cores and some other models are highly unlikely, (2) the most promising cluster is a trimer-plus- monomer type of distorted cubane, and (3) S* = 5/2 excited states are higher than the first excited state with S* = 3/2 (majority) or S* = 1/2 (minority).
The photosynthetic oxidation of water to oxygen occurs in photosystem II (PSII) at an active site composed of a tetranuclear cluster of manganese ions, a redox active tyrosine, YZ, and two essential cofactors, calcium and chloride. Recently, several experimental observations have led to the proposal of a metalloradical catalytic cycle in which water oxidation occurs via hydrogen-atom abstraction by the tyrosyl radical from water bound to the manganese cluster. This model predicts a close proximity between the radical tyrosine, Yz•, and the Mn cluster and the involvement of the radical in a bifurcated hydrogen bond. Magnetic resonance techniques have been used in this work to probe the interaction of the tyrosyl radical with its environment in PSII samples in which the catalytic cycle is blocked by acetate treatment and the enzyme is trapped in a paramagnetic S2Yz• state. Radical interaction with the metal cluster has been studied via simulations of the EPR spectra obtained for this state. The simulations were based on a radical-pair model and included terms for both electron−electron dipolar and exchange interactions. The results show a dominant exchange interaction between the radical and the manganese cluster in these preparations and led to an estimate of 8−9 Å for the spin−spin distance. ESEEM spectroscopy and 1H2O/2H2O exchange were used to study interactions of the S2Yz• state with exchangeable hydrogen nuclei in the site. Two-pulse ESEEM data show features expected for a radical-pair species, in support of our analysis of the continuous-wave EPR spectrum. An ESEEM analysis based on an electron spin 1/2, nuclear spin 1 model shows that both two- and three-pulse ESEEM data are consistent with four deuterons that exhibit an electron−nuclear dipole−dipole coupling of 0.42 MHz. The validity of this analysis and its implications for the oxygen-evolving apparatus are discussed.
Cross-reaction rate constants k12 (22 °C) at pH 7.0 have been determined for the reduction of FeIII2 and tyrosyl-radical-containing active-R2 from E. coli ribonucleotide reductase with eight organic radicals (OR), e.g., MV•+ from methyl viologen. The more reactive OR's were generated in situ using pulse radiolysis (PR) techniques, and other OR's were generated by prior reduction of the parent with dithionite, followed by stopped-flow (SF) studies. In both procedures it was necessary to include consideration of doubly-reduced parent forms. Values of k12 are in the range 109 to 104 M-1 s-1 and reduction potentials Eo1 for the OR vary from −0.446 to +0.194 V. Samples of E. coli active-R2 also have an FeIII2 met-R2 component (with no Tyr•), which in the present work was close to 40%. From separate experiments met-R2 gave similar k12 rate constants (on average 66% bigger) to those for active-R2, suggesting that reduction of the FeIII2 center is the common rate-limiting step. A single Marcus free-energy plot of log k12 − 0.5 log f vs −Eo1/0.059 describes all the data, and the slope of 0.54 is in satisfactory agreement with the theoretical value of 0.50. It is concluded that the rate-limiting step involves electron transfer. In addition, the intercept at −Eo1/0.059 = 0 is 5.94, where values of the reduction potential and self-exchange rate constant for met-R2 contribute to this value. To maintain electroneutrality at the 10 Å buried active site H+ uptake is also required. For both e- and H+ transfer the conserved pathway Trp-48, Asp-237, His-118 to FeA is a possible candidate requiring further examination.
The O2-evolving complex (OEC) of photosystem II (PSII) contains a tetramanganese (Mn4) cluster, a redox-active tyrosine, and Ca2+/Cl- ions, but its molecular structure has not been determined. Vibrational spectroscopy has the potential of providing new structural information for the OEC, particularly the Mn4 cluster. Toward this goal, the vibrational characteristics of the OEC of PSII were examined using near-infrared (NIR) excitation Raman spectroscopy. NIR excitation decreases the background contribution from chlorophyll emission/Raman scattering and affords the opportunity of probing selectively low-energy electronic transitions of the Mn4 cluster. The primary emphasis of the Raman study was on the low-frequency range of the spectrum (220−620 cm-1) where Mn−ligand vibrational modes are expected to occur. The low-frequency region was examined for both the S1 and S2 oxidation states of the Mn4 cluster. A particular effort was made to probe a NIR transition of the S2 state that has been reported to mediate photoconversion from the multiline to the g = 4.1 form of the S2 state [Boussac et al. Biochemistry 1996, 35, 6984−6989]. The Raman studies revealed the following: (1) the Raman spectra of Mn-depleted PSII and PSII in the S2 state are nearly identical; (2) the Raman spectrum of PSII in the S1 state displays several unique low-frequency bands not present in the S2 state that can be assigned as Mn−ligand vibrational modes and appear to maximize in intensity at λex 820 nm; and (3) several of the S1 state Raman bands are shifted by D2O/H2O exchange. Collectively, these results indicate that the S1 state of the Mn4 cluster (1) has a NIR electronic transition from which resonance enhanced Raman scattering can be induced and (2) is coordinated by at least two H2O or OH- groups. The studies reported herein also demonstrate the potential of NIR-excitation Raman techniques for probing selectively the OEC in PSII and, in particular, for characterizing the coordination environment of the Mn4 cluster.
The signature feature of the enzymatic cycle of the peroxidase family of metabolizing heme proteins is formation of the catalytically active compound I species from the inactive ferric resting form, via a putative transient peroxide bound intermediate. While there is some evidence for this intermediate, the mechanism of formation of compound I from it and the role of nearby amino acids in facilitating it are still unresolved. To further probe this mechanism and investigate the possible role of the protein in compound I formation, molecular dynamics simulations of the peroxide bound complex of horseradish peroxidase isoenzyme C (HRP-C−HOOH) were performed. For such a typical peroxidase, a role of two conserved amino acids in the distal binding pocket, histidine and arginine, has been suggested in facilitating the peroxide O−O bond cleavage necessary for compound I formation. Since HRP functions cover a wide range of pH values, protein simulations were carried out for two models differing only in the state of protonation of the conserved histidine. The neutral histidine corresponds to a high-pH model, and the cationic histidine corresponds to a low-pH model. The unique robust H bonds identified in the molecular dynamics simulations of the two models suggest two different modes of binding of the peroxide to the heme iron, different mechanisms of compound I formation, and a different role for the key HRP residues involved in its formation in the two models.
The oxygen-evolving complex of Photosystem II in plants and cyanobacteria catalyzes the oxidation of two water molecules to one molecule of dioxygen. A tetranuclear Mn complex is believed to cycle through five intermediate states (S0−S4) to couple the four-electron oxidation of water with the one-electron photochemistry occurring at the Photosystem II reaction center. We have used X-ray absorption spectroscopy to study the local structure of the Mn complex and have proposed a model for it, based on studies of the Mn K-edges and the extended X-ray absorption fine structure of the S1 and S2 states. The proposed model consists of two di-μ-oxo-bridged binuclear Mn units with Mn−Mn distances of 2.7 Å that are linked to each other by a mono-μ-oxo bridge with a Mn−Mn separation of 3.3 Å. The Mn−Mn distances are invariant in the native S1 and S2 states. This report describes the application of X-ray absorption spectroscopy to S3 samples created under physiological conditions with saturating flash illumination. Significant changes are observed in the Mn−Mn distances in the S3 state compared to the S1 and the S2 states. The two 2.7 Å Mn−Mn distances that characterize the di-μ-oxo centers in the S1 and S2 states are lengthened to 2.8 and 3.0 Å in the S3 state, respectively. The 3.3 Å Mn−Mn and Mn−Ca distances also increase by 0.04−0.2 Å. These changes in Mn−Mn distances are interpreted as consequences of the onset of substrate/water oxidation in the S3 state. Mn-centered oxidation is evident during the S0→S1 and S1→S2 transitions. We propose that the changes in Mn−Mn distances during the S2→S3 transition are the result of ligand or water oxidation, leading to the formation of an oxyl radical intermediate formed at a bridging or terminal position. The reaction of the oxyl radical with OH-, H2O, or an oxo group during the subsequent S state conversion is proposed to lead to the formation of the O−O bond. Models that can account for changes in the Mn−Mn distances in the S3 state and the implications for the mechanism of water oxidation are discussed.
Spin state considerations are proposed to sharply limit the possible O−O bond-forming steps in water oxidation by the oxygen evolving center of Photosystem II. A series of intermediates are proposed for the Kok S states on the basis of quantum chemical studies on simple model complexes; these are also consistent with the main biophysical data. Only one Mn atom in the active site cluster is thought to be redox-active and mediate O−O bond formation. A key concept is the formation of an unreactive MnO oxo at the S2 state, followed by its conversion to a reactive Mn−O• oxyl form at the S3 level, with radical character on the oxyl oxygen, at which point O−O bond formation can occur by a coupling between the oxyl and an outer-sphere water molecule. An MnOOH intermediate at S3 is proposed to lose a hydrogen atom to give O2. The role of the Ca cofactor is to bring about a 5- to 6-coordination change at S2, necessary for formation of a reactive oxo in S3. The chloride cofactor is assigned the role of charge neutralization.
The evaluation of Mn X-ray absorption fine structure (EXAFS) studies on the oxygen-evolving complex (OEC) from photosystem II is described for preparations from both spinach and the cyanobacterium Synechococcus sp. poised in the S[sub 1] and S[sub 2] states. In addition to reproducing previous results suggesting the presence of bis([mu]-oxo)-bridged Mn centers in the OEC, a Fourier transform peak due to scatterers at an average distance of > 3 [angstrom] is detected in both types of preparation. In addition, subtle but reproducible changes are found in the relative amplitudes of the Fourier transform peaks due to mainly O ([approximately]1.8 [angstrom]) and Mn ([approximately] 2.7 [angstrom]) neighbors upon cryogenic advance from the S[sub 1] to the S[sub 2] state. Analysis of the peak due to scatterers at [approximately] 3 [angstrom] favors assignment to (per 4 Mn in the OEC) 1-2 heavy atom (Mn, Ca) scatterers at an average distance of 3.3-3.4 [angstrom]. The EXAFS data of several multinuclear Mn model compounds containing such scattering interactions are analyzed and compared with the data for the OEC. Structural models for the OEC are evaluated on the basis of these results. 40 refs., 9 figs., 5 tabs.
The structural and electronic consequences of reduction and oxidation of a peroxo-bridged Mn2IV/IV dimer, Mn2(μ-O)2(μ-O2)(NH3)62+, are examined using approximate density functional theory. In both cases, the initial electron-transfer step is localized on the metal centers, but subsequent structural rearrangement results in transfer of the excess charge to the μ-O2 unit, with concomitant regeneration of the Mn2IV/IV core. Two-electron reduction results in population of the O−O σ* orbital and complete cleavage of the O−O bond, whereas two-electron oxidation depopulates the O−O π* orbital, forming molecular oxygen. The coupling between the metal centers (antiferromagnetic or ferromagnetic) affects the stability of the intermediate species, in which the redox process is metal based, and hence influences the kinetic barrier to bond formation or cleavage. Reductive cleavage of the O−O σ bond is favored when the metal centers are antiferromagnetically coupled, whereas oxidative formation of the π component of the O−O bond is favored by ferromagnetic coupling. The possible implications for the relationship between structure and function in the oxygen-evolving complex found in photosynthetic organisms are discussed.
Photoactive yellow protein (PYP) undergoes a light-driven cycle of color and protonation states that is part of a mechanism of bacterial phototaxis. This article concerns functionally important protonation states of PYP and the interactions that stabilize them, and changes in the protonation state during the photocycle. In particular, the chromophore pKa is known to be shifted down so that the chromophore is negatively charged in the ground state (dark state) even though it is buried in the protein, while nearby Glu46 has an unusually high pKa. The photocycle involves changes of one or both of these protonation states. Calculations of pKa values and protonation states using a semi-macroscopic electrostatic model are presented for the wild-type and three mutants, in both the ground state and the bleached (I2) intermediate state. Calculations allowing multiple H-bonding arrangements around the chromophore also have been carried out. In addition, ground-state pKa values of the chromophore have been measured by UV−visible spectroscopy for the wild-type and the same three mutants. Because of the unusual protonation states and strong electrostatic interactions, PYP represents a severe test of the ability of theoretical models to yield correct calculations of electrostatic interactions in proteins. Good agreement between experiment and theory can be obtained for the ground state provided the protein interior is assumed to have a relatively low dielectric constant, but only partial agreement between theory and experiment is obtained for the bleached state. We also present a reinterpretation of previously published data on the pH-dependence of the recovery of the ground state from the bleached state. The new analysis implies a pKa value of 6.37 for Glu46 in the bleached state, which is consistent with other available experimental data, including data that only became available after this analysis. The new analysis suggests that signal transduction is modulated by the titration properties of the bleached state, which are in turn determined by electrostatic interactions. Overall, the results of this study provide a quantitative picture of the interactions responsible for the unusual protonation states of the chromophore and Glu46, and of protonation changes upon bleaching.
The temperature dependence of donor side reactions was analysed within the framework of the Marcus theory of nonadiabatic
electron transfer. The following results were obtained for PS II membrane fragments from spinach: (1) the reorganisation energy
of P680+• reduction by YZ is of the order of 0.5 eV in samples with a functionally fully competent water oxidising complex (WOC); (2) destruction of
the WOC by Tris-washing gives rise to a drastic increase of λ to values of the order of 1.6 eV; (3) the reorganisation energies
of the oxidation steps in the WOC are dependent, on the redox states S
i
with values of about 0.6 eV for the reactions YZ
OX
S
0→YZ
S
1 and YZ
OX
S
1→YZ
S
2, 1.6 eV for the reaction YZ
OX
S
2→YZ
S
3 and 1.1 eV (above a characteristic temperature uc of about 6 °C) for the reaction YZ
OX
S
3→→YZ
S
0+O2. Using an empirical rate constant-distance relationship, the van der Waals distance between YZ and P680 was found to be about 10 Å, independent of the presence or absence of the WOC, whereas the distance between YZ and the manganese cluster in the WOC was ≥15 Å. Based on the calculated activation energies the environment of YZ is inferred to be almost "dry" and hydrophobic when the WOC is intact but becomes enriched with water molecules after WOC
destruction. Furthermore, it is concluded that the transition S
2→S
3 is an electron transfer reaction gated by a conformational change, i.e. it comprises significant structural changes of functional
relevance. Measurements of kinetic H/D isotope exchange effects support the idea that none of these reactions is gated by
the break of a covalent O-H bond. The implications of these findings for the mechanism of water oxidation are discussed.
The stabilization of the photosynthetic charge separation in Photosystem II by secondary reactions was studied using chlorophyll luminescence induced by electric field pulses in a suspension of preilluminated osmotically swollen chloroplasts. This ‘electroluminescence’ was measured as a function of the delay time between illumination and field pulse, and as a function of the number of preilluminating flashes. The result is a survey of, in principle, all stabilization and deactivation processes beyond the state Z+Q−A, which is formed within the approx. 20 μs time resolution of the method. Most of these could be identified with known secondary electron transfer reactions. A 20-fold stabilization with a half-time of 330 μs is attributed to Q−A reoxidation. No further stabilization at the acceptor side seemed to occur and no flash number dependence was detected, although a normal QB/Q−B oscillation was found in ultraviolet absorbance. With regard to the donor side, the data are consistent with the known S-state-dependent Z+ reduction times and indicate values of 9, 5 and 65 for the equilibrium constants associated with this reaction on the transitions S1→S2, S2→S3 and S3→S0(O2) respectively. Z+ reduction was found to be electrogenic and exposed to about 5% of the membrane potential. An 0.1 s phase in S0 is attributed to oxygen release. S2 and S3 are further stabilized in two phases of unknown origin with half-times of 15 ms and 0.4 s, followed by a final 20 s phase attributed to deactivation. In S1, Z+ reduction was probably hidden in an unresolved fast phase present on all transitions, but in addition a 350 μs phase was found, which might be related to proton release. In nearly 20% of the Photosystem II reaction centers electron transfer beyond Q−A was inhibited. In these centers Z+ reduction appeared to take about 1 ms and charge recombination followed in phases of about 8, 80 and 800 ms half-time.