Secondary Quinone Acceptor Research Articles

Effect of different LED-lighting quality conditions on growth and photosynthetic characteristics of saffron plants (Crocus sativus L.)

The effects of different light-emitting diode (LED) lights on saffron growth and photosynthetic characteristic were explored. Physiological mechanisms were explained by chlorophyll a fluorescence transient curves (OJIP) and JIP-test parameters. A decrease in the red to blue light ratio resulted in negative effects, particularly for monochromatic blue (B) LED light; saffron seedlings showed reduced chlorophyll accumulation, inhibited leaf elongation, and decreased photosynthetic performance. In the OJIP curve, the higher positive K-band observed for B LED light indicated that oxygen-evolving complex activation significantly decreased. B LED light inhibited the electron transport between primary quinone acceptor and secondary quinone acceptor as well as the existence of reducing plastoquinone centers, and increased energy dissipation of reaction centers. Otherwise, the red to blue light ratio of 2:1 had a positive effect on saffron cultivation, resulting in the longest leaf lengths, highest chlorophyll content, and photosynthetic characteristics. This study provides theoretical guidance for saffron agricultural practices.

Exogenously spermidine alleviates damage from drought stress in the photosystem II of tall fescue

Drought stress is one of the major limiting factors to crop productivity around the globe. It has been well documented that spermidine (Spd) plays an important key role in plant growth and development, especially in the defense response to stress. The objective of this study was to explore the effect of Spd on protecting photosynthetic apparatus in tall fescue under drought stress. Spd application significantly improved the OJIP (fluorescence rise kinetics O-J-I-P) curve compared to non-Spd application during drought. Exogenous Spd exhibited higher F<sub>J</sub> (fluorescence value at the J-step (2 ms) of OJIP) and F<sub>P</sub> (maximal recorded fluorescence intensity, at the peak P of OJIP) than non-Spd application. Moreover, normalised total complementary area (S<sub>m</sub>) and the number of Q<sub>A</sub> (primary quinone acceptor of PS II) reduction events (N) significantly reduced after the application of Spd in tall fescue under drought stress. In terms of quantum yields and efficiencies and specific energy fluxes, exogenous Spd notably decreased the values of efficiency of electron transfer from Q<sub>B</sub> (secondary quinone acceptor of PS II) to PSI acceptors (δR<sub>0</sub>), absorption flux per RC (ABS/RC) and trapping flux per RC (TR<sub>0</sub>/RC) compared to the non-Spd application without watering. All the above suggested that exogenous Spd facilitated the photosynthetic system of tall fescue in drought. These observations involved in the electron transport capacity of photosystem II assist in understanding better the protective role of exogenous Spd in tall fescue under drought stress.

Responses of the photosynthetic apparatus of Abies koreana to drought under different light conditions

AbstractThe threat of drought to trees is predicted to increase due to global warming. In a forest stand, the physiological responses of trees can differ depending on the light conditions. We analyzed photosynthetic rate, photopigment, and chlorophyll a fluorescence transient (OJIP test) of Abies koreana E.H. Wilson, under different light (full sunlight and 35 and 75% shading) and water conditions (well‐watering, W; and no‐watering, NW) to examine the combined effect of light and water. After 21 days of no‐watering, we observed decreases in the photosynthetic rate and photopigment contents and quality, impairment of electron transfer from primary to secondary quinone acceptor, inactivation of reaction center, and lower photosynthetic performance index, especially under full sunlight. The time required for recovery after re‐watering was also slower under full sunlight. In conclusion, the adverse effects of drought on light absorption and utility of A. koreana in the photosynthetic process were much greater under high light intensity compared to shading conditions, which alleviated these effects.

Effect of different methods of Ca2+ extraction from PSII oxygen-evolving complex on the QA- oxidation kinetics.

Lumenal extrinsic proteins PsbO, PsbP, and PsbQ of photosystem II (PSII) protect the catalytic cluster Mn4CaO5 of oxygen-evolving complex (OEC) from the bulk solution and from soluble compounds in the surrounding medium. Extraction of PsbP and PsbQ proteins by NaCl-washing together with chelator EGTA is followed also by the depletion of Ca2+ cation from OEC. In this study, the effects of PsbP and PsbQ proteins, as well as Ca2+ extraction from OEC on the kinetics of the reduced primary electron acceptor (QA-) oxidation, have been studied by fluorescence decay kinetics measurements in PSII membrane fragments. We found that in addition to the impairment of OEC, removal of PsbP and PsbQ significantly slows the rate of electron transfer from QA- to the secondary quinone acceptor QB. Electron transfer from QA- to QB in photosystem II membranes with an occupied QB site was slowed down by a factor of 8. However, addition of EGTA or CaCl2 to NaCl-washed PSII did not change the kinetics of fluorescence decay. Moreover, the kinetics of QA- oxidation by QB in Ca-depleted PSII membranes obtained by treatment with citrate buffer at pH 3.0 (such treatment keeps all extrinsic proteins in PSII but extracts Ca2+ from OEC) was not changed. The results obtained indicate that the effect of NaCl-washing on the QA- to QB electron transport is due to PsbP and PsbQ extrinsic proteins extraction, but not due to Ca2+ depletion.

Kinetic modeling of electron transfer reactions in photosystem I complexes of various structures with substituted quinone acceptors.

The reduction kinetics of the photo-oxidized primary electron donor P700 in photosystem I (PS I) complexes from cyanobacteria Synechocystis sp. PCC 6803 were analyzed within the kinetic model, which considers electron transfer (ET) reactions between P700, secondary quinone acceptor A1, iron-sulfur clusters and external electron donor and acceptors-methylviologen (MV), 2,3-dichloro-naphthoquinone (Cl2NQ) and oxygen. PS I complexes containing various quinones in the A1-binding site (phylloquinone PhQ, plastoquinone-9 PQ and Cl2NQ) as well as F X-core complexes, depleted of terminal iron-sulfur F A/F B clusters, were studied. The acceleration of charge recombination in F X-core complexes by PhQ/PQ substitution indicates that backward ET from the iron-sulfur clusters involves quinone in the A1-binding site. The kinetic parameters of ET reactions were obtained by global fitting of the P700+ reduction with the kinetic model. The free energy gap ΔG 0 between F X and F A/F B clusters was estimated as -130meV. The driving force of ET from A1 to F X was determined as -50 and -220meV for PhQ in the A and B cofactor branches, respectively. For PQ in A1A-site, this reaction was found to be endergonic (ΔG 0 = +75meV). The interaction of PS I with external acceptors was quantitatively described in terms of Michaelis-Menten kinetics. The second-order rate constants of ET from F A/F B, F X and Cl2NQ in the A1-site of PS I to external acceptors were estimated. The side production of superoxide radical in the A1-site by oxygen reduction via the Mehler reaction might comprise ≥0.3% of the total electron flow in PS I.

Catecholamine-functionalized graphene as a biomimetic redox shuttle for solar water oxidation.

In natural photosynthesis, solar energy is converted to chemical energy through a cascaded, photoinduced charge transfer chain that consists of primary and secondary acceptor quinones (i.e., QA and QB). This leads to an exceptionally high near-unity quantum yield. Inspired by the unique multistep architecture of charge transfer in nature, we have synthesized a catecholamine-functionalized, reduced graphene oxide (RGO) film as a redox mediator that can mimic quinone acceptors in photosystem II. We used polynorepinephrine (PNE) as a redox-shuttling chemical. We also used it to coat graphene oxide (GO) and to reduce GO to RGO. The quinone ligands in PNE, which are characterized by a charge transfer involving two electrons and two protons, acted as electron acceptors that facilitated charge transfer in photocatalytic water oxidation. Furthermore, PNE-coated RGO film promoted fast charge separation in [Ru(bpy)3]2+ and increased the activity of cobalt phosphate on photocatalytic water oxidation more than two-fold. The results suggest that our bio-inspired strategy for the construction of a forward charge transfer pathway can provide more opportunities to realize efficient artificial photosynthesis.

The Acceptor Side of Photosystem II Is the Initial Target of Nitrite Stress in Synechocystis sp. Strain PCC 6803.

Nitrite, a common form of inorganic nitrogen (N), can be used as a nitrogen source through N assimilation. However, high levels of nitrite depress photosynthesis in various organisms. In this study, we investigated which components of the photosynthetic electron transfer chain are targeted by nitrite stress in Synechocystis sp. strain PCC 6803 cells. Measurements of whole-chain and photosystem II (PSII)-mediated electron transport activities revealed that high levels of nitrite primarily impair electron flow in PSII. Changes in PSII activity in response to nitrite stress occurred in two distinct phases. During the first phase, which occurred in the first 3 h of nitrite treatment, electron transfer from the primary quinone acceptor (QA) to the secondary quinone acceptor (QB) was retarded, as indicated by chlorophyll (Chl) a fluorescence induction, S-state distribution, and QA- reoxidation tests. In the second phase, which occurred after 6 h of nitrite exposure, the reaction center was inactivated and the donor side of photosystem II was inhibited, as revealed by changes in Chl fluorescence parameters and thermoluminescence and by immunoblot analysis. Our data suggest that nitrite stress is highly damaging to PSII and disrupts PSII activity by a stepwise mechanism in which the acceptor side is the initial target. IMPORTANCE In our previous studies, an alga-based technology was proposed to fix the large amounts of nitrite that are released from NOX-rich flue gases and proved to be a promising industrial strategy for flue gas NOX bioremediation (W. Chen et al., Environ Sci Technol 50:1620-1627, 2016, https://doi.org/10.1021/acs.est.5b04696; X. Zhang et al., Environ Sci Technol 48:10497-10504, 2014, https://doi.org/10.1021/es5013824). However, the toxic effects of high concentrations of nitrite on algal cells remain obscure. The analysis of growth rates, photochemistry, and protein profiles in our study provides important evidence that the inhibition by nitrite occurs in two phases: in the first phase, electron transfer between QA- and QB is retarded, whereas in the second, the donor side of PSII is affected. This is an excellent example of investigating the "early" inhibitory effects (i.e., within the first 6 h) on the PSII electron transfer chain in vivo This paper provides novel insights into the mechanisms of nitrite inhibition of photosynthesis in an oxygenic phototrophic cyanobacterium.

Effect of ionic strength on intra-protein electron transfer reactions: The case study of charge recombination within the bacterial reaction center

It is a common believe that intra-protein electron transfer (ET) involving reactants and products that are overall electroneutral are not influenced by the ions of the surrounding solution. The results presented here show an electrostatic coupling between the ionic atmosphere surrounding a membrane protein (the reaction center (RC) from the photosynthetic bacterium Rhodobacter sphaeroides) and two very different intra-protein ET processes taking place within it. Specifically we have studied the effect of salt concentration on: i) the kinetics of the charge recombination between the reduced primary quinone acceptor QA− and the primary photoxidized donor P+; ii) the thermodynamic equilibrium (QA−↔QB−) for the ET between QA− and the secondary quinone acceptor QB. A distinctive point of this investigation is that reactants and products are overall electroneutral. The protein electrostatics has been described adopting the lowest level of complexity sufficient to grasp the experimental phenomenology and the impact of salt on the relative free energy level of reactants and products has been evaluated according to suitable thermodynamic cycles. The ionic strength effect was found to be independent on the ion nature for P+QA− charge recombination where the leading electrostatic term was the dipole moment. In the case of the QA−↔QB− equilibrium, the relative stability of QA− and QB− was found to depend on the salt concentration in a fashion that is different for chaotropic and kosmotropic ions. In such a case both dipole moment and quadrupole moments of the RC must be considered.

Chlorophyll a fluorescence of typical desert plant Alhagi sparsifolia Shap. at two light levels

Alhagi sparsifolia Shap. is exposed to a high-irradiance environment as the main vegetation found in the forelands of the Taklamakan Desert. We investigated chlorophyll a fluorescence emission of A. sparsifolia seedlings grown under ambient (HL) and shade (LL) conditions. Our results indicated that the fluorescence intensity in the leaves was significantly higher for LL-grown plants than that under HL. High values of the maximum quantum yield of PSII for primary photochemistry (φPo) and the quantum yield that an electron moves further than QA - (φEo) in the plants under LL conditions suggested that the electron flow from QA - (primary quinone electron acceptors of PSII) to QB (secondary quinone acceptor of PSII) or QB - was enhanced at LL compared to natural HL conditions. The efficiency/probability with which an electron from the intersystem electron carriers was transferred to reduce end electron acceptors at the PSI acceptor side and the quantum yield for the reduction of end electron acceptors at the PSI acceptor side were opposite to φPo, and φEo. Thus, we concluded that the electron transport on the donor side of PSII was blocked under LL conditions, while acceptor side was inhibited at the HL conditions. The PSII activity of electron transport in the plants grown in shade was enhanced, while the energy transport from PSII to PSI was blocked compared to the plants grown at HL conditions. Furthermore, PSII activity under HL was seriously affected in midday, while the plants grown in shade enhanced their energy transport.

Hydrogen bond network around the semiquinone of the secondary quinone acceptor Q(B) in bacterial photosynthetic reaction centers.

By utilizing a combined pulsed EPR and DFT approach, the high-resolution structure of the QB site semiquinone (SQB) was determined. The development of such a technique is crucial toward an understanding of protein-bound semiquinones on the structural level, as (i) membrane protein crystallography typically results in low resolution structures, and (ii) obtaining protein crystals in the semiquinone form is rarely feasible. The SQB hydrogen bond network was investigated with Q- (∼34 GHz) and X-band (∼9.7 GHz) pulsed EPR spectroscopy on fully deuterated reactions centers from Rhodobacter sphaeroides. Simulations in the SQB g-tensor reference frame provided the principal values and directions of the H-bond proton hyperfine tensors. Three protons were detected, one with an anisotropic tensor component, T = 4.6 MHz, assigned to the histidine NδH of His-L190, and two others with similar anisotropic constants T = 3.2 and 3.0 MHz assigned to the peptide NpH of Gly-L225 and Ile-L224, respectively. Despite the strong similarity in the peptide couplings, all hyperfine tensors were resolved in the Q-band ENDOR spectra. The Euler angles describing the series of rotations that bring the hyperfine tensors into the SQB g-tensor reference frame were obtained by least-squares fitting of the spectral simulations to the ENDOR data. These Euler angles show the locations of the hydrogen bonded protons with respect to the semiquinone. Our geometry optimized model of SQB used in previous DFT work is in strong agreement with the angular constraints from the spectral simulations, providing the foundation for future joint pulsed EPR and DFT semiquinone structural determinations in other proteins.

A Model of Light-Activated Changes in Reaction Centers of Purple Bacteria

We have studied the slow dynamics of isolated complexes of chlorophyll-containing membrane proteins of photosynthetic Reaction Centers (RCs) of Rb. sphaeroides R-26 induced by light fluxes. The analysis of a variation of the absorption of a solution of RCs in the frame of the three-level model is carried out. The equation defining the ratio of the populations of the electron levels of the primary and secondary quinones is deduced. The solution of this equation has non-Boltzmann character even in the stationary case and depends on the exciting light intensity. A model of the dynamics of levels of RC which accounts the presence of polarization processes in the vicinity of the secondary quinone acceptor (QB) is proposed. It is assumed that all RCs have the same structure but can be in different states, whose characteristics depend on both the time interval having passed after the absorption of a light quantum and the local viscosity and elasticity of the environment of QB. The presence of deformational properties of the environment of QB yields the possibility for a configuration of RC to be changed, which agrees with the experimental data.

Long-range interaction between the Mn4CaO5 cluster and the non-heme iron center in photosystem II as revealed by FTIR spectroelectrochemistry.

It is known that inactivation of the Mn4CaO5 cluster, the catalytic center of water oxidation in photosystem II (PSII), induces a positive shift of the redox potential (Em) of the primary quinone electron acceptor QA by ∼+150 mV, resulting in suppression of the electron transfer from QA to the secondary quinone acceptor QB. Although the relevance of this Em(QA(-)/QA) shift to the photoprotection of PSII has been debated, its molecular mechanism is still enigmatic from a structural viewpoint because QA is ∼40 Å from the Mn4CaO5 cluster. In this work, we have investigated the influence of Mn depletion on the Em of the non-heme iron, which is located between QA and QB, and its surrounding structure. Electrochemical measurements in combination with Fourier transform infrared (FTIR) spectroscopy revealed that Mn depletion shifts Em(Fe(2+)/Fe(3+)) by +18 mV, which is ∼8 times smaller than the shift of Em(QA(-)/QA). Comparison of the Fe(2+)/Fe(3+) FTIR difference spectra between intact and Mn-depleted PSII samples showed that Mn depletion altered the pKa's of a His ligand to the non-heme iron, most probably the D1-His215 interacting QB, and a carboxylate group, possibly D1-Glu244, coupled with the non-heme iron. It was further shown that Mn depletion influences the C≡N vibration of bromoxynil bound to the QB site, indicative of the modification of the QB binding site. On the basis of these results, we discuss the mechanism of a long-range interaction between the donor and acceptor sides of PSII.

Cationic screening of charged surface groups (carboxylates) affects electron transfer steps in photosystem-II water oxidation and quinone reduction

The functional or regulatory role of long-distance interactions between protein surface and interior represents an insufficiently understood aspect of protein function. Cationic screening of surface charges determines the morphology of thylakoid membrane stacks. We show that it also influences directly the light-driven reactions in the interior of photosystem II (PSII). After laser-flash excitation of PSII membrane particles from spinach, time courses of the delayed recombination fluorescence (10μs–10ms) and the variable chlorophyll-fluorescence yield (100μs–1s) were recorded in the presence of chloride salts. At low salt-concentrations, a stimulating effect was observed for the S-state transition efficiency, the time constant of O2-formation at the Mn4Ca-complex of PSII, and the halftime of re-oxidation of the primary quinone acceptor (Qa) by the secondary quinone acceptor (Qb). The cation valence determined the half-effect concentrations of the stimulating salt effect, which were around 6μM, 200μM and 10mM for trivalent (LaCl3), bivalent (MgCl2, CaCl2), and monovalent cations (NaCl, KCl), respectively. A depressing high-salt effect also depended strongly on the cation valence (onset concentrations around 2mM, 50mM, and 500mM). These salt effects are proposed to originate from electrostatic screening of negatively charged carboxylate sidechains, which are found in the form of carboxylate clusters at the solvent-exposed protein surface. We conclude that the influence of electrostatic screening by solvent cations manifests a functionally relevant long-distance interaction between protein surface and electron-transfer reactions in the protein interior. A relation to regulation and adaptation in response to environmental changes is conceivable.

Differentiation between two strains of microalga Parachlorella kessleri using modern spectroscopic method.

BackgroundThe differentiation between wild type of Parachlorella kessleri and its mutant strains PC Mut2, PC Mut4 by using the Multi-functional Plant Efficiency Analyzer (М-РЕА-2) was studied. Mutant algal cells of P. kessleri have been obtained by UV-C during 3 and 10 min respectively.ResultsLight-induced kinetics of prompt fluorescence (OJIP transients), delayed fluorescence and modulated reflection at 820 nm (redox transitions of P700 in PSI) showed disturbance of electron transport flow in photosystem II (PSII) and an increase fraction of non-reducing centers of secondary quinone acceptors of electron (QB). In addition, the amplitudes of the fast and slow peak in the kinetics of the delayed light emission and non-photochemical fluorescence quenching ( NPQ) were significantly reduced in mutant cells, indicating low level of the membrane energization of photosynthetic membranes. Changes of photosynthetic reactions of mutants may lead to an increase of the carotenoids content, which protect cells against the light stress.ConclusionIt is suggested to use parameters of induction curves of prompt and delayed fluorescence to characterize mutant algal cells in biotechnological studies.Electronic supplementary materialThe online version of this article (doi:10.1186/s40529-014-0053-7) contains supplementary material, which is available to authorized users.

The spectral-kinetic indicators of relaxation processes following the electron stabilization into the acceptor compartment of photosynthetic RCs of bacteria.

Longterm temporary stabilization of the electron in the acceptor part of the photosynthetic reaction center (RC) is essential for subsequent efficient trans� fer of reducing equivalents to the photosynthetic membrane. Comparative study of the kinetics of redox reactions of photoactive bacteriochlorophyll of the RC of purple bacteria and quinone acceptors in their indi� vidual absorption bands is an informative approach to study the mechanisms of this stabilization. The analy� sis of the revealed kinetic differences makes it possible to estimate the activation energy and the characteristic time of the transition relaxation process associated with the stabilization of the electron on the final quinone acceptor and establish the hydrogen bonds in the quinone environment that are involved in this pro� cess. Conformational dynamics of protein-pigment complexes of photosynthetic RCs plays the crucial role in all stages of light energy conversion, including the temporary stabilization of electrons in the quinone acceptor component of the RCs of purple bacteria (1, 2). The important functional role of effective oper� ation of this component of the electron transport chain (ETC) is determined primarily by the fact that it couples the very fast initial stages of the lightinduced charge separation with the significantly slower diffu� sioncontrolled reactions of transferring the reducing equivalents to the membrane. Ultimately, two quinone molecules, QA and QB, integrated into the RC struc� ture, are actively involved in the coupled electron- proton transport processes that lead to the formation of a proton gradient across the photosynthetic mem� brane, required for the synthesis of ATP. These two quinones have different redox potentials, providing the driving force for the vector electron transfer from the primary to the secondary quinone acceptor. Since

Acetate in mixotrophic growth medium affects photosystem II in Chlamydomonas reinhardtii and protects against photoinhibition

Chlamydomonas reinhardtii is a photoautotrophic green alga, which can be grown mixotrophically in acetate-supplemented media (Tris–acetate–phosphate). We show that acetate has a direct effect on photosystem II (PSII). As a consequence, Tris–acetate–phosphate-grown mixotrophic C. reinhardtii cultures are less susceptible to photoinhibition than photoautotrophic cultures when subjected to high light. Spin-trapping electron paramagnetic resonance spectroscopy showed that thylakoids from mixotrophic C. reinhardtii produced less 1O2 than those from photoautotrophic cultures. The same was observed in vivo by measuring DanePy oxalate fluorescence quenching. Photoinhibition can be induced by the production of 1O2 originating from charge recombination events in photosystem II, which are governed by the midpoint potentials (Em) of the quinone electron acceptors. Thermoluminescence indicated that the Em of the primary quinone acceptor (QA/QA−) of mixotrophic cells was stabilised while the Em of the secondary quinone acceptor (QB/QB−) was destabilised, therefore favouring direct non-radiative charge recombination events that do not lead to 1O2 production. Acetate treatment of photosystem II-enriched membrane fragments from spinach led to the same thermoluminescence shifts as observed in C. reinhardtii, showing that acetate exhibits a direct effect on photosystem II independent from the metabolic state of a cell. A change in the environment of the non-heme iron of acetate-treated photosystem II particles was detected by low temperature electron paramagnetic resonance spectroscopy. We hypothesise that acetate replaces the bicarbonate associated to the non-heme iron and changes the environment of QA and QB affecting photosystem II charge recombination events and photoinhibition.

PSII photochemistry in vegetative buds and needles of Norway spruce (Picea abiesL. Karst.) probed by OJIP chlorophyllafluorescence measurement

Vegetative buds represent developmental stage of Norway spruce (Picea abies L. Karst.) needles where chloroplast biogenesis and photosynthetic activity begin. We used the analyses of polyphasic chlorophyll a fluorescence rise (OJIP) to compare photosystem II (PSII) functioning in vegetative buds and fully photosynthetically active mature current-year needles. Considerably decreased performance index (PIABS) in vegetative buds compared to needles pointed to their low photosynthetic efficiency. Maximum quantum yield of PSII (Fv/Fm) in buds was slightly decreased but above limited value for functionality indicating that primary photochemistry of PSII is not holdback of vegetative buds photosynthetic activity. The most significant difference observed between investigated developmental stages was accumulation of reduced primary quinine acceptor of PSII (QA-) in vegetative buds, as a result of its limited re-oxidation by passing electrons to secondary quinone acceptor, QB. We suggest that reduced electron transfer from QA- to QB could be the major limiting factor of photosynthesis in vegetative buds.

A High Redox Potential Form of Cytochrome c550 in Photosystem II from Thermosynechococcus elongatus

Cytochrome c(550) (cyt c(550)) is a component of photosystem II (PSII) from cyanobacteria, red algae, and some other eukaryotic algae. Its physiological role remains unclear. In the present work, measurements of the midpoint redox potential (E(m)) were performed using intact PSII core complexes preparations from a histidine-tagged PSII mutant strain of the thermophilic cyanobacterium Thermosynechococcus (T.) elongatus. When redox titrations were done in the absence of redox mediators, an E(m) value of +200 mV was obtained for cyt c(550). This value is ∼300 mV more positive than that previously measured in the presence of mediators (E(m) = -80 mV). The shift from the high potential form (E(m) = +200 mV) to the low potential form (E(m) = -80 mV) of cyt c(550) is attributed to conformational changes, triggered by the reduction of a component of PSII that is sequestered and out of equilibrium with the medium, most likely the Mn(4)Ca cluster. This reduction can occur when reduced low potential redox mediators are present or under highly reducing conditions even in the absence of mediators. Based on these observations, it is suggested that the E(m) of +200 mV obtained without mediators could be the physiological redox potential of the cyt c(550) in PSII. This value opens the possibility of a redox function for cyt c(550) in PSII.

High-Resolution Two-Dimensional1H and14N Hyperfine Sublevel Correlation Spectroscopy of the Primary Quinone of Photosystem II

Quinones are naturally occurring isoprenoids that are widely exploited by photosynthetic reaction centers. Protein interactions modify the properties of quinones such that similar quinone species can perform diverse functions in reaction centers. Both type I and type II (oxygenic and nonoxygenic, respectively) reaction centers contain quinone cofactors that serve very different functions as the redox potential of similar quinones can operate at up to 800 mV lower reduction potential when present in type I reaction centers. However, the factors that determine quinone function in energy transduction remain unclear. It is thought that the location of the quinone cofactor, the geometry of its binding site, and the "smart" matrix effects from the surrounding protein environment greatly influence the functional properties of quinones. Photosystem II offers a unique system for the investigation of the factors that influence quinone function in energy transduction. It contains identical plastoquinones in the primary and secondary quinone acceptor sites, Q(A) and Q(B), which exhibit very different functional properties. This study is focused on elucidating the tuning and control of the primary semiquinone state, Q(A)(-), of photosystem II. We utilize high-resolution two-dimensional hyperfine sublevel correlation spectroscopy to directly probe the strength and orientation of the hydrogen bonds of the Q(A)(-) state with the surrounding protein environment of photosystem II. We observe two asymmetric hydrogen bonding interactions of reduced Q(A)(-) in which the strength of each hydrogen bond is affected by the relative nonplanarity of the bond. This study confirms the importance of hydrogen bonds in the redox tuning of the primary semiquinone state of photosystem II.

Engineering of an alternative electron transfer path in photosystem II

The initial steps of oxygenic photosynthetic electron transfer occur within photosystem II, an intricate pigment/protein transmembrane complex. Light-driven electron transfer occurs within a multistep pathway that is efficiently insulated from competing electron transfer pathways. The heart of the electron transfer system, composed of six linearly coupled redox active cofactors that enable electron transfer from water to the secondary quinone acceptor Q(B), is mainly embedded within two proteins called D1 and D2. We have identified a site in silico, poised in the vicinity of the Q(A) intermediate quinone acceptor, which could serve as a potential binding site for redox active proteins. Here we show that modification of Lysine 238 of the D1 protein to glutamic acid (Glu) in the cyanobacterium Synechocystis sp. PCC 6803, results in a strain that grows photautotrophically. The Glu thylakoid membranes are able to perform light-dependent reduction of exogenous cytochrome c with water as the electron donor. Cytochrome c photoreduction by the Glu mutant was also shown to significantly protect the D1 protein from photodamage when isolated thylakoid membranes were illuminated. We have therefore engineered a novel electron transfer pathway from water to a soluble protein electron carrier without harming the normal function of photosystem II.