The mechanism of photosynthetic water oxidation

Photosynthetie water oxidation is unique to plants and cyanobacteria, it occurs in thylakoid membranes. The components associated with this process include: a reaction center polypeptide, having a molecular weight (Mr) of 47–50 kilodaltons (kDa), containing a reaction center chlorophyll a labeled as P680, a plastoquinol(?)-electron donor Z, a primary electron acceptor pheophytin, and a quinone electron acceptor Q_A; three ‘extrinsic’ polypeptides having Mr of approximately 17 kDa, 23 kDa, and 33 kDa; and, in all likelihood, an approximately 34 kDa ‘intrinsic’ polypeptide associated with manganese (Mn) atoms. In addition, chloride and calcium ions appear to be essential components for water oxidation. Photons, absorbed by the so-called photosystem II, provide the necessary energy for the chemical oxidation-reduction at P680; the oxidized P680 (P680⁺), then, oxidizes Z, which then oxidizes the water-manganese system contained, perhaps, in a protein matrix. The oxidation of water, leading to O₂ evolution and H⁺ release, requires four such independent acts, i.e., there is a charge accumulating device (the so-called S-states). In this minireview, we have presented our current understanding of the reaction center P680, the chemical nature of Z, a possible working model for water oxidation, and the possible roles of manganese atoms, chloride ions, and the various polypeptides, mentioned above. A comparison with cytochrome c oxidase, which is involved in the opposite process of the reduction of O₂ to H₂O, is stressed. This minireview is a prelude to the several minireviews, scheduled to be published in the forthcoming issues of Photosynthesis Research, including those on photosystem II (by H.J. van Gorkom); polypeptides of the O₂-evolving system (by D.F. Ghanotakis and C.F. Yocum); and the role of chloride in O₂ evolution (by S. Izawa).     

Cyclic electron transfer in photosystem II in the marine diatom Phaeodactylum tricornutum

In Phaeodactylum tricornutum Photosystem II is unusually resistant to damage by exposure to high light intensities. Not only is the capacity to dissipate excess excitations in the antenna much larger and induced more rapidly than in other organisms, but in addition an electron transfer cycle in the reaction center appears to prevent oxidative damage when secondary electron transport cannot keep up with the rate of charge separations. Such cyclic electron transfer had been inferred from oxygen measurements suggesting that some of its intermediates can be reduced in the dark and can subsequently compete with water as an electron donor to Photosystem II upon illumination. Here, the proposed activation of cyclic electron transfer by illumination is confirmed and shown to require only a second. On the other hand the dark reduction of its intermediates, specifically of tyrosine Y_D, the only Photosystem II component known to compete with water oxidation, is ruled out. It appears that the cyclic electron transfer pathway can be fully opened by reduction of the plastoquinone pool in the dark. Oxygen evolution reappears after partial oxidation of the pool by Photosystem I, but the pool itself is not involved in cyclic electron transfer.     

光合作用氧释放机理研究进展

植物在光合作用过程中不仅为同化CO2提供能量和还原力,同时裂解水放出氧气。放氧反应主要由光系统Ⅱ(PSⅡ)氧化侧的4个锰原子组成的锰簇催化完成的。因此,锰簇在光合放氧过程中起看至关重要的作用。文章概述了对锰簇及其微环境的结构和功能的研究进展。     

Early Archean origin of Photosystem II

Photosystem II is a photochemical reaction center that catalyzes the light‐driven oxidation of water to molecular oxygen. Water oxidation is the distinctive photochemical reaction that permitted the evolution of oxygenic photosynthesis and the eventual rise of eukaryotes. At what point during the history of life an ancestral photosystem evolved the capacity to oxidize water still remains unknown. Here, we study the evolution of the core reaction center proteins of Photosystem II using sequence and structural comparisons in combination with Bayesian relaxed molecular clocks. Our results indicate that a homodimeric photosystem with sufficient oxidizing power to split water had already appeared in the early Archean about a billion years before the most recent common ancestor of all described Cyanobacteria capable of oxygenic photosynthesis, and well before the diversification of some of the known groups of anoxygenic photosynthetic bacteria. Based on a structural and functional rationale, we hypothesize that this early Archean photosystem was capable of water oxidation to oxygen and had already evolved protection mechanisms against the formation of reactive oxygen species. This would place primordial forms of oxygenic photosynthesis at a very early stage in the evolutionary history of life.     

Turnover of the D1 protein and of Photosystem II in a Synechocystis 6803 mutant lacking Tyrz

The Photosystem II reaction center is rapidly inactivated by light, particularly at higher light intensity. One of the possible factors causing this phenomenon is the oxidized primary donor, P680⁺, which may be harmful to Photosystem II because of its highly oxidizing nature. However, no direct evidence specificially implicating P680⁺ in photoinhibition has been obtained yet. To investigate whether P680⁺ is harmful to Photosystem II, turnover of the D1 protein and of the Photosystem II reaction center complex were measured in vivo in a mutant of the cyanobacterium Synechocystis sp. PCC 6803, in which the physiological donor to P680⁺, Tyr_z, was genetically deleted. In this mutant, D1 degradation in the light is an order of magnitude faster than in wild type. The most straightforward explanation of this phenomenon is that accumulation of P680⁺ leads to an increased rate of turnover of the Photosystem II reaction center complex, which is compatible with the hypothesis of destructive oxidation by P680⁺ that is damaging to the Photosystem II complex.     

Water cleavage by solar radiation—an inspiring challenge of photosynthesis research

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 S_i (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 S_i, 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 Y_Z ^ox 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.Abbreviations b559 cytochrome b559 - BChl bacteriochlorophyll - Chl chlorophyll - CP47 Chl a containing a 47 kDa polypeptide - D1/D2 polypeptides of the PS II reaction center - ENDOR electron nuclear double resonance - EPR electron paramagnetic resonance - ESEEM electron spin echo envelope modulation - EXAFS extended X-ray absorption fine structure - FTIR Fourier transform infrared - NMR nuclear magnetic resonance - P680, P700 photoactive Chl a of PS II and PS I, respectively - PS II Photosystem II - Q_A special plastoquinone of PS II - S_i redox states of WOC - WOC water oxidizing complex - WOS water oxidizing site - UV/VIS ultraviolet/visible - Y_D, Y_Z redox active tyrosines of polypeptides D2 and D1, respectively     

EPR characteristics of oxygen-evoloving Photosytem II particles from Phormidium laminosum

The EPR characteristics of oxygen evolving particles prepared from Phormidium laminosum are described. These particles are enriched in Photosystem II allowing EPR investigation of signals which were previously small or masked by those from Photosystem I in other preparations. EPR signals from a Signal II species and high potential cytochrome b-559 appear as they are photooxidised at cryogenic temperatures by Photosystem II. The Signal II species is a donor close to the Photosystem II reaction centre and may represent part of the charge accumulation system of water oxidation. An EPR signal from an iron-sulphur centre which may represent an unidentified component of photosynthetic electron transport is also described.The properties of the oxygen evolving particles show that the preparation is superior to chloroplasts or unfractionated algal membranes for the study of Photosystem II with a functional water oxidation system.     

Sequence-specific 1H, 13C, and 15N backbone assignment of Psb27 from Synechocystis PCC 6803

Photosystem II (PSII) is a large membrane protein complex that uses light to split water into molecular oxygen, protons, and electrons. Here we report the ¹H, ¹⁵N and ¹³C backbone chemical shift assignments for the Psb27 protein of Photosystem II from Synechocystis PCC 6803. These assignments will now provide the basis for the structural analysis of the Psb27 protein.     

Towards artificial photosynthesis: ruthenium-manganese chemistry mimicking photosystem II reactions

The reaction center Photosystem II is a key component of the most successful solar energy converting machinery on earth: the oxygenic photosynthesis. Photosystem II uses light to drive the reduction of plastoquinone and the oxidation of water. Water-oxidation is catalyzed by a manganese cluster and gives the organism an abundant source of electrons. The principles of photosynthesis have inspired chemists to mimic these reactions in artificial molecular assemblies. Synthetic light-harvesting antennae and light-induced charge separation systems have been demonstrated by several groups. More recently, there has been an increasing effort to mimic Photosystem II by coupling light-driven charge separation to water oxidation, catalyzed by synthetic manganese complexes.     

Photosystem II in a mutant of Chlamydomonas reinhardtii lacking the 23 kDa psbP protein shows increased sensitivity to photoinhibition in the absence of chloride

The psbP gene product, the so called 23 kDa extrinsic protein, is involved in water oxidation carried out by Photosystem II. However, the protein is not absolutely required for water oxidation. Here we have studied Photosystem II mediated electron transfer in a mutant of Chlamydomonas reinhardtii, the FUD 39 mutant, that lacks the psbP protein. When grown in dim light the Photosystem II content in thylakoid membranes of FUD 39 is approximately similar to that in the wild-type. The oxygen evolution is dependent on the presence of chloride as a cofactor, which activates the water oxidation with a dissociation constant of about 4 mM. In the mutant, the oxygen evolution is very sensitive to photoinhibition when assayed at low chloride concentrations while chloride protects against photoinhibition with a dissociation constant of about 5 mM. The photoinhibition is irreversible as oxygen evolution cannot be restored by the addition of chloride to inhibited samples. In addition the inhibition seems to be targeted primarily to the Mn-cluster in Photosystem II as the electron transfer through the remaining part of Photosystem II is photoinhibited with slower kinetics. Thus, this mutant provides an experimental system in which effects of photoinhibition induced by lesions at the donor side of Photosystem II can be studied in vivo.Abbreviations Chl chlorophyll - DCIP 2,6-dichlorophenolindophenol - DPC 2,2-diphenylcarbonic dihydrazide - HEPES 4-(2-hydroxyethyl)-1-piperazinethanesulfonic acid - P₆₈₀ the primary electron donor to PS II - PpBQ phenyl-p-benzoquinone - PS II Photosystem II - Q_A the first quinone acceptor of PS II - Q_B the second quinone acceptor of PS II - SDS sodium dodecyl sulfate - Tris tris(hydroxymethyl)aminomethane - Tyr_D accessory electron donor on the D₂-protein - Tyr_Z tyrosine residue, acting as electron carrier between P₆₈₀ and the water oxidizing system     

Quality Control of Photosystem II: Lipid Peroxidation Accelerates Photoinhibition under Excessive Illumination

Environmental stresses lower the efficiency of photosynthesis and sometimes cause irreversible damage to plant functions. When spinach thylakoids and Photosystem II membranes were illuminated with excessive visible light (100–1,000 µmol photons m⁻¹ s⁻¹) for 10 min at either 20°C or 30°C, the optimum quantum yield of Photosystem II decreased as the light intensity and temperature increased. Reactive oxygen species and endogenous cationic radicals produced through a photochemical reaction at and/or near the reaction center have been implicated in the damage to the D1 protein. Here we present evidence that lipid peroxidation induced by the illumination is involved in the damage to the D1 protein and the subunits of the light-harvesting complex of Photosystem II. This is reasoned from the results that considerable lipid peroxidation occurred in the thylakoids in the light, and that lipoxygenase externally added in the dark induced inhibition of Photosystem II activity in the thylakoids, production of singlet oxygen, which was monitored by electron paramagnetic resonance spin trapping, and damage to the D1 protein, in parallel with lipid peroxidation. Modification of the subunits of the light-harvesting complex of Photosystem II by malondialdehyde as well as oxidation of the subunits was also observed. We suggest that mainly singlet oxygen formed through lipid peroxidation under light stress participates in damaging the Photosystem II subunits.     

Evidence of singlet oxygen evolution by whole living cells of Chlamydomonas reinhardtii

The oxygen evolved by Chlamydomonas reinhardtii in the light is measured simultaneously with a Clark electrode and with the nitrosodimethylaniline-imidazole colorimetric method which is specific for singlet oxygen. Experiments with wild-type and FuD7 mutant cells (unable to synthesize the D1 protein of Photosystem II), with dichlorophenyldimethylurea (which blocks electron transfer from Photosystem II to Photosystem I) and with dibromothymoquinone (which diverts electrons from their normal path between the two photosystems), as well as with hydroxylamine (an inactivator of the water-splitting part of Photosystem II and a competitor of water for electron donation to it), all point to the dependence of detected singlet oxygen on photolysis of water by Photosystem II.Abbreviations DBMIB Dibromothymoquinone - DCMU Dichlorophenyldimethylurea - PS I and PS II Photosystems I and II - RNO para-nitrosodimethylaniline Contribution of the Centre interdisciplinaire de Biochimie de Oxygène.     

Involvement of the donor tyrosine-D1 (Yd) in Photosystem II electron transport in the green alga, Chlamydobotrys stellata

The light-induced oxidation of the accessory donor tyrosine-D (Y_D) has been studied by measurements of the EPR Signal II_slow at room temperature in the autotrophically and photoheterotrophically cultivated alga Chlamydobotrys stellata. After illumination and dark adaptation, Y_D Signal II_slow was observed only in autotrophic algae, i.e. under conditions of a linear photosynthetic electron transfer from water to NADP⁺. The addition of artificial electron acceptors phenyl-p-benzoquinone (PPQ) or dichloro-p-benzoquinone (DCQ) to the autotrophic cells caused an almost negligible increase of this signal. When photosynthetic electron flow and oxygen evolution were diminished by removal of the carbon source CO₂ and addition of acetate (photoheterotrophy), a pronounced Y_D Signal II_slow was seen only in presence of DCQ or PPQ. Several possibilities are discussed to explain the absence of Y_D Signal II_slow in photoheterotrophic Chl. stellata such as the existence of a cyclic PS II electron flow very effectively reducing P₆₈₀ and thereby preventing the possibility of Y_D oxidation. Artificial electron acceptors withdraw electrons from this cycle thus keeping the primary quinone acceptor, Q_A, oxidized and thereby diminishing the reduction of P₆₈₀ ⁺ by cyclic PSII. This leads to the appearance of the Y_D Signal II_slow also in the photoheterotrophically grown algae.Abbreviations A-band- thermoluminescence band associated with S₂Q_A ^- charge recombination - DCQ- 2,5-dichlorobenzoquinone - D₂- structure protein of Photosystem II - EPR- electron paramagnetic resonance - OEC- oxygen evolving complex - PPQ- phenyl-p-benzoquinone - PS II- Photosystem II - P₆₈₀- reaction center of Photosystem II - Q-band- thermoluminescence band associated with S₂Q_A ^- charge recombination - S_i- oxidation levels of the OEC - Y_D- tyrosine-D accessory donor to P₆₈₀ - Y_Z- tyrosine-Z electron donor to P₆₈₀ Dedicated to Prof. Dr E. Schnepf/Heidelberg.     

Proton release during the redox cycle of the water oxidase

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 s) and before the oxidation of the Mn-cluster by Y_z ⁺ is completed. By these electrostatically driven reactions the catalytic center accumulates bases. An additional slow phase is observed during the oxygen evolving step, S₃S₄S₀. 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.Abbreviations BBY-membranes Photosystem II-enriched membrane fragments prepared after Berthold, Babcock and Yocum (1981) - BSA bovine serum albumin - Chl chlorophyll - CAB-protein chlorophyll a/b-binding protein - core particles oxygen evolving reaction center core particles of Photosystem II - Cyt cytochrome - DCBQ 2,5-dichloro-p-benzoquinone - IML intermittent light - P-680 primary electron donor of Photosystem II - PS II Photosystem II - Y_z tyrosine residue on the D1 polypeptide, electron carrier between manganese and P-680 - photochemical reaction      

Light saturation curves show competence of the water splitting complex in inactive Photosystem II reaction centers

Photosystem II complexes of higher plants are structurally and functionally heterogeneous. While the only clearly defined structural difference is that Photosystem II reaction centers are served by two distinct antenna sizes, several types of functional heterogeneity have been demonstrated. Among these is the observation that in dark-adapted leaves of spinach and pea, over 30% of the Photosystem II reaction centers are unable to reduce plastoquinone to plastoquinol at physiologically meaningful rates. Several lines of evidence show that the impaired reaction centers are effectively inactive, because the rate of oxidation of the primary quinone acceptor, Q_A, is 1000 times slower than in normally active reaction centers. However, there are conflicting opinions and data over whether inactive Photosystem II complexes are capable of oxidizing water in the presence of certain artificial electron acceptors. In the present study we investigated whether inactive Photosystem II complexes have a functional water oxidizing system in spinach thylakoid membranes by measuring the flash yield of water oxidation products as a function of flash intensity. At low flash energies (less that 10% saturation), selected to minimize double turnovers of reaction centers, we found that in the presence of the artificial quinone acceptor, dichlorobenzoquinone (DCBQ), the yield of proton release was enhanced 20±2% over that observed in the presence of dimethylbenzoquinone (DMBQ). We argue that the extra proton release is from the normally inactive Photosystem II reaction centers that have been activated in the presence of DCBQ, demonstrating their capacity to oxidize water in repetitive flashes, as concluded by Graan and Ort (Biochim Biophys Acta (1986) 852: 320–330). The light saturation curves indicate that the effective antenna size of inactive reaction centers is 55±12% the size of active Photosystem II centers. Comparison of the light saturation dependence of steady state oxygen evolution in the presence of DCBQ or DMBQ support the conclusion that inactive Photosystem II complexes have a functional water oxidation system.Abbreviations DCBQ 2,6-dichloro-p-benzoquinone - DMBQ 2,5-dimethyl-p-benzoquinone - F_o initial fluorescence level using dark-adapted thylakoids - Inactive reaction centers reaction centers inactive in plastoquinone reduction - PS II Photosystem II - Q_A primary quinone acceptor of Photosystem II - Q_B secondary quinone acceptor of Photosystem II Department of Plant Biology, University of IllinoisDepartment of Physiology & Biophysics, University of Illinois     

Polypeptides of chloroplastic and cytoplastic origin required for development of Photosystem II activity,and chlorophyll-protein complexes,in Euglena gracilis Z chloroplast membranes

The development of photosynthetic activity and synthesis of chloroplast membrane polypeptides was studied during greening of Euglena gracilis Z in alternate light-dark-light cycles. The results show: (a) The development of both Photosystem II and Photosystem I can be dissociated from chlorophyll synthesis. (b) Most of the polypeptides required for development of Photosystem I are already synthesized during the initial light period (10–12 h); the further rise in Photosystem I activity in the dark is not inhibited by cycloheximide nor by chloramphenicol. (c) The development of Photosystem II requires continuous de novo synthesis of polypeptides and is inhibited by chloramphenicol. The water-splitting activity already present at the end of the first light period decays in the presence of chloramphenicol while that of 1,5-diphenylcarbazide oxidation is only partially retained. The activity can be repaired in the absence of chlorophyll synthesis and is correlated with the de novo synthesis of polypeptides of 50 000–60 000 daltons. The synthesis of these polypeptides and associated repair of Photosystem II activity is not inhibited by cycloheximide. (d) The chloroplast membranes can be resolved into about 40 distinct polypeptides, among them several in the molecular weight range 50 000–60 000, 20 000–35 000 and 10 000–15 000, which are major membrane constitutents. (e) The synthesis of two major polypeptides (M_r = 20 000–30 000) required for the formation of chlorophyll-protein complex(es) containing chlorophyll a and traces of chlorophyll b (CPII?) is light-dependent and cycloheximide-inhibited. It is concluded that the synthesis and addition to the growing membrane of chlorophyll and polypeptides required for the formation of Photosystem II and Photosystem I complexes can be dissociated in time. The H₂O-splitting enzyme(s) and possibly other components of Photosystem II complex are of chloroplastic origin and turn over in the dark while at least some of the chlorophyll binding polypeptides are of cytoplastic origin and their synthesis is light-controlled.     

Photooxidation of ferrocyanide and iodide ions and associated phosphorylation in NH2OH-treated chloroplasts

NH₂OH-treated, non-water oxidizing chloroplasts are shown to be capable of oxidizing ferrocyanide and I^? via Photosystem II at appreciable rates (? 200 μequiv/h per mg chlorophyll). Using methylviologen as electron acceptor, ferrocyanide oxidation can be measured as O₂ uptake, as ferricyanide formation, or as H⁺ consumption (2 Fe²⁺ + 2H⁺ + O₂ → 2 Fe³⁺ + H₂O₂). I^? oxidation can be measured as methylviologen-mediated O₂ uptake, or spectrophotometrically, using ferricyanide as electron acceptor. The oxidation product I₂ is re-reduced, as it is formed, by unknown reducing substances in the reaction system.The rate-saturating concentrations of these donors are very high: 30 mM with ferricyanide and 15 mM with I^?. Relatively lipophilic Photosystem II donors such as catechol, benzidine and p-aminophenol saturate the photooxidation rate at much lower concentrations (< 0.5 mM). It thus seems that the oxidation of hydrophilic reductants such as ferricyanide and I^? is limited by permeability barriers. Very likely the site of Photosystem II oxidation is embedded in the thylakoid membrane or is situated on the inner surface of the membrane.The efficiency of phosphorylation (P/e₂) is 0.5 to 0.6 with ferrocyanide and about 0.5 with I^?. In contrast the P/e₂ ratio is 1.0 to 1.2 when water, catechol, p-aminophenol or benzidine serves as electron donor. These differences imply that only one of two phosphorylation sites operate when ferrocyanide and I^? are oxidized. Ferrocyanide and I^? are also chemically distinct from other Photosystem II donors in that their oxidation does not involve proton release. It is suggested that the mechanism of energy conservation associated with Photosystem II may be only operative when the removal of electrons from the donor results in release of protons (i.e. with water, hydroquinones, phenylamines, etc.).     

Partial characterization of the biosynthesis and integration of the Photosystem II reaction centers in the thylakoid membrane of Chlamydomonas reinhardtii

Pulse-labeling of wild-type and a Photosystem II mutant strain of Chlamydomonas reinhardtii was carried out in the presence or absence of inhibitors of either cytoplasmic or chloroplast ribosomes, and their thylakoid membrane polypeptides were analyzed by polyacrylamide gel electrophoresis. A pulse-chase study was also done on the wild-type strain in the presence of anisomycin, an inhibitor of protein synthesis on cytoplasmic ribosomes. The following results were obtained: the Photosystem II reaction center is mainly composed of integral membrane proteins synthesized within the chloroplast. Several of the proteins of the Photosystem II reaction center are post-translationally modified, after they have been inserted in the thylakoid membrane.     

Activating anions that replace Cl- in the O2-evolving complex of photosystem II slow the kinetics of the terminal step in water oxidation and destabilize the S2 and S3 states

Photosystem II, the multisubunit protein complex that oxidizes water to O2, requires the inorganic cofactors Ca2+ and Cl- to exhibit optimal activity. Chloride can be replaced functionally by a small number of anionic cofactors (Br-, NO3-, NO2-, I-), but among these anions, only Br- is capable of restoring rates of oxygen evolution comparable to those observed with Cl-. UV absorption difference spectroscopy was utilized in the experiments described here as a probe to monitor donor side reactions in photosystem II in the presence of Cl- or surrogate anions. The rate of the final step of the water oxidation cycle was found to depend on the activating anion bound at the Cl- site, but the kinetics of this step did not limit the light-saturated rate of oxygen evolution. Instead, the lower oxygen evolution rates supported by surrogate anions appeared to be correlated with an instability of the higher oxidation states of the oxygen-evolving complex that was induced by addition of these anions. Reduction of these states takes place not only with I- but also with NO2- and to a lesser extent even with NO3- and Br- and is not related to the ability of these anions to bind at the Cl- binding site. Rather, it appears that these anions can attack higher oxidation states of the oxygen evolving complex from a second site that is not shielded by the extrinsic 17 and 23 kDa polypeptides and cause a one-electron reduction. The decrease of the oxygen evolution rate may result from accumulated damage to the reaction center protein by the one-electron oxidation product of the anion.     

Calcium ions can be substituted for the 24-kDa polypeptide in photosynthetic oxygen evolution

Photosystem II particles were prepared from spinach chloroplasts with Triton X-100, and treated with 1.0 M NaCl to remove polypeptides of 24 kDa and 18 kDa and to reduce the photosynthetic oxygen-evolution activity by about half. Oxygen-evolution activity was restored almost to the original level with 10 mM Ca²⁺, in a similar manner to the rebinding of 24-kDa polypeptide. Other cations such as magnesium, sodium and manganese ions could not restore any oxygen-evolution activity. These observations, together with a kinetic analysis, suggest that Ca²⁺ can be substituted for the 24-kDa polypeptide in photosynthetic oxygen evolution in Photosystem II particles.