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Plant Physiology and Biochemistry xxx (2014) 1e13

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Plant Physiology and Biochemistry

journal homepage: www.elsevier .com/locate/plaphy

Review

A nano-sized manganese oxide in a protein matrix as a naturalwater-oxidizing siteq

Mohammad Mahdi Najafpour a,b,**, Mohadeseh Zarei Ghobadi a, Behzad Haghighi a,b,Tatsuya Tomo c,d, Robert Carpentier e, Jian-Ren Shen f, Suleyman I. Allakhverdiev g,h,*

aDepartment of Chemistry, Institute for Advanced Studies in Basic Sciences (IASBS), Zanjan 45137-66731, IranbCenter of Climate Change and Global Warming, Institute for Advanced Studies in Basic Sciences (IASBS), Zanjan 45137-66731, IrancDepartment of Biology, Faculty of Science, Tokyo University of Science, Kagurazaka 1-3, Shinjuku-ku, Tokyo 162-8601, Japand PRESTO, Japan Science and Technology Agency (JST), Saitama 332-0012, JapaneDepartement de Chimie Biochimie et Physique, Université du Québec à Trois Rivières, C.P. 500, Québec G9A 5H7, CanadafGraduate School of Natural Science and Technology, Faculty of Science, Okayama University, Okayama 700-8530, Japang Institute of Plant Physiology, Russian Academy of Sciences, Botanicheskaya Street 35, Moscow 127276, Russiah Institute of Basic Biological Problems, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia

a r t i c l e i n f o

Article history:Received 5 December 2013Accepted 26 January 2014Available online xxx

Keywords:H2 productionPhotosystem IINano-sized MneCa clusterWater-oxidizing centerOxygen evolutionArtificial photosynthesisNano-layered Mn oxide

q This article is dedicated to the memory of Dr. WJanuary 10, 2014.* Corresponding author. Institute of Basic Biologica

of Sciences, Pushchino, Moscow Region 142290, Rufax: þ7 496 7330 532.** Corresponding author. Department of Chemistry, Iin Basic Sciences (IASBS), Zanjan 45137-66731, Irafax: þ98 241 415 3232.

E-mail addresses: mmnajafpour@iasbs.ac.ir (Mallakhverdiev@gmail.com, sallakhverdiev@yandex.ru

http://dx.doi.org/10.1016/j.plaphy.2014.01.0200981-9428/� 2014 Elsevier Masson SAS. All rights re

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a b s t r a c t

The purpose of this review is to present recent advances in the structural and functional studies of water-oxidizing center of Photosystem II and its surrounding protein matrix in order to synthesize artificialcatalysts for production of clean and efficient hydrogen fuel.

� 2014 Elsevier Masson SAS. All rights reserved.

1. Introduction

Nowadays, one of the significant threats facing mankind is theclimate change caused by the increased CO2 production associatedwith the consumption of fossil fuels. The aftermath of this globalwarming problem is the reduction of food sources because of un-favorable conditions for plants growth. Hence, active research forthe production of clean and affordable energy is needed (Barber andTran, 2012). To this end, nature has been an inspiration wellspring.

arwick Hillier who died on

l Problems, Russian Academyssia. Tel.: þ7 496 7731 837;

nstitute for Advanced Studiesn. Tel.: þ98 241 415 3201;

.M. Najafpour), suleyman.(S.I. Allakhverdiev).

served.

, M.M., et al., A nano-sized mttp://dx.doi.org/10.1016/j.pla

Photosynthesis is a process used by green plant, algae,and cyanobacteria to convert carbon dioxide and into organicmolecules and oxygen in the presence of light. Despite numerousproposals for the origin of photosynthesis on Earth, there is nodetailed information to support any of them (Olson andBlankenship, 2004). Most researchers agree that nonoxygenicphotosynthesis performed approximately 3.2e3.5 billion yearsago. Also, the evidence from rock fossils indicates that the greatoxidation event (GOE) occurred from 2.48 billion years ago, coin-ciding with the evolution of cyanobacteria (Konhauser et al., 2011;Govindjee and Shevela, 2011). Probably, the first experimentexploring the nature of photosynthesis was performed by vanHelmont in 1648 (see Williams, 1957). Now, we have extensiveinformation about the details of this phenomenon. Photosynthesisis a two-step process: light reactions in the grana and CO2 fixationin the stroma of the chloroplast. The light dependent part is ful-filled by two sequential photosystems (photosystem II (PSII) andphotosystem I (PSI)).

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The first step in the PSII (membraneeprotein complex) is thelight-induced excitation of chlorophyll a (P680) and loss one elec-tron. P680þ compensates its electron deficient state by the donation ofan electron from the amino acid tyrosine (Yz) and subsequently, Yz

þ

is rereduced by an electron obtained from water cleavage into O2,electrons andHþ owing to the catalytic action of thewater-oxidizingcenter known as the WOC (or oxygen-evolving center (OEC))(McEvoy and Brudvig, 2006; Najafpour, 2006; Najafpour et al.,2012a). The electron lost by P680 passes through a series of elec-tron carriers, then is trapped by another chlorophyll a (P700) in thePSI. P700 also loses an electron following light-induced excitationthat will be used to produce NADPH. This series of electron transferswill also lead to ATP synthesis. The second step of photosynthesisknown as the Calvin Cycle can take place in the absence of light. Inthis process, the atmospheric carbon dioxide is captured and con-verted to carbohydrate by the use of the energy from the NADPHand ATP produced in the first stage (Golbeck, 2006). The scrutiny ofthe photosynthesis process can help the researchers to find acompound with high efficiency and stability in artificial systems forwater splitting (Eq. (3)), water oxidation (Eq. (1)) and reduction (Eq.(2)), that can also be used to store renewable energies (Xu et al.,2009; Jiao and Frei, 2010; Najafpour, 2011; Najafpour et al., 2011).However, in water splitting, water oxidation is a bottleneck.

4H2O / 4Hþ þ O2þ4e (1)

4Hþ þ 4e / 2H2 (2)

2H2O / 2H2 þ O2 (3)

Such reactions (Eqs. (1)e(3)) can occur in both artificial andnatural systems. In this review, we will highlight the importanceand the structure of the WOC in PSII. Furthermore, we will discuss

Fig. 1. The S-state cycle showing how the absorption of four photons of light (hn) by P680 dseries of five intermediates (S0, S1, S2, S3 and S4). Protons (Hþ) are released during this cycleaided by the redox active tyrosine YZ. Also shown are half-times for the various steps of the c(2008) by Elsevier.

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various proposed mechanisms for the WOC in the nature andsynthetic compounds.

2. Water-oxidizing center: position, significance, andstructure

One of the most important reactions in nature is the photosyn-thetic water oxidation. As a result, oxygen is evolvedwith a turnoverof up to 100e400 released O2molecules per second (Dismukes et al.,2009), which is the main source of the atmosphere’s oxygen. Also,this process is a major inspiration source for scientists to synthesizenew compounds in view of the production of clean hydrogen fuel.The main focus is the reaction center of the WOC that includes aMn4O5Ca cluster which is surrounded by a protein environmentmanaging reaction coordinates, proton motion, and water availabil-ity. The four-electron oxidation mechanism of water oxidation withan undermost activation energy is as follows (Wiechen et al., 2012).

From the thermodynamic point of view, four-electron wateroxidation is most likely to occur because other possible reactionssuch as four-sequential one-electron oxidation or two-sequentialtwo-electron oxidation are more energy-consuming. Each oxida-tion state of the WOC is known as an “S-state”, in which the oxida-tion level gradually increases from S0 to S4 (Fig. 1) (Dau andHaumann, 2008; Barber, 2002). The WOC couple the sequentialone-electron reduction of P680 to the four electrons oxidation ofwater. All the S-state transitions, except the S4 / S0, are induced bythe photochemical oxidation of chlorophyll (P680þ ), which oxidizesthe WOC via a redox-active tyrosine. Oxygen evolution occurs in thelight independent transition of S4 / S0 (McEvoy and Brudvig, 2006).

In this section, we briefly go over the history of the structuralinvestigation of the Mn cluster. The first idea about Mn involve-ment in photosynthesis was introduced in 1937 by Pirson (1937).About 40 years later, Jaklevic et al. recorded the X-ray absorption

rives the splitting of two water molecules and formation of O2 through a consecutiveexcept for the S1 to S2 transition. Electron donation from the Mn4Ca cluster to P680þ is

ycle (Barber, 2002). Image was reprinted with permission from Barber (2002) Copyright

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spectrum of the Mn in a leaf (Jaklevic et al., 1977). They estimated aMn concentration of 10e50 ppm in the chloroplast of green plants.Continuous studies conducted by M. P. Klein et al. revealed that: i)the presence of di-m-oxo bridged pairs of Mn atoms in the WOC byEXAFS analysis (Kirby et al., 1981); ii) the attendance of Mn withoxidation state higher than þ2 via analysis of the X-ray absorptionedges (Kirby et al., 1981). By observing the variations in X-rayabsorption edge of Mn in PSII, Yachandra group concluded that Mnpartaked in the light-driven electron-transfer and was oxidized asthe WOC advances from the S1 to S2 state (Goodin et al., 1984). V. K.Yachandra et al., in 1986 surveyed the K-edge spectra and EXAFS ofMn complex. It was deduced that the oxidation state of Mnincreased in the transition from S0 to S1 and S1 to S2 but no changeoccurred from S2 to S3. In addition, by observation of similaritybetween the structural analysis results of the complexes fromspinach and Synechococcus, they suggested the basic structure ofthe Mn center was conserved over the two billion years (Yachandraet al., 1986). Therefore, the bridgedMn complex was proposed as anessential feature of the O2 evolving complex. The attendances oftwo inequivalent di-m-oxo bridged binuclear structures in the S3state were suggested by Guiles in 1990 (Guiles et al., 1990).

Fig. 2. Structure of PSII with assignment of protein subunits and cofactors. Arrangement ofcompletely, with part of the second monomer related by the local-C2 axis (filled ellipse on thThe view direction is from the luminal side, perpendicular to the membrane plane. The a-antennae, and CP43 and CP47 by circles. Seven unassigned a-helices are shown in grey. The fo(Zouni et al., 2001). Image was reprinted with permission from Zouni et al. (2001) Copyrig

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Sauer and his colleague offered the dimer of dimers modelwhich assumed the Mn complexes composed of two Mn “dimers”.Several structural models including 3 þ 1 arrangement of Mnatoms in the cluster were also investigated based on the EXAFSdata by DeRose et al. (1994). This structure was confirmed bymagnetic resonance experiments (Peloquin et al., 2000), andtheoretical studies (Siegbahn, 2000). The first detailed structurestudy of PSII was reported by Witt and Saenger (Zouni et al., 2001).They isolated dimeric PSII from the thermophilic cyanobacteriumand depicted the three-dimensional structure of WOC at 3.8 �Aresolution (Fig. 2). The identified cluster structure was introducedas follows: three Mn ions located in the corners of an isoscelestriangle accompany with a fourth Mn ion near the center of thetriangle. Nobuo Kamiya and Jian-Ren Shen in 2003 analyzed thePSII which was extracted from Thermosynechococcus vulcanus(Kamiya and Shen, 2003). They confirmed the previous providedstructure with a difference that all four Mn atoms were placedapproximately in the same plane. J. Barber and S. Iwata in 2004suggested a cubane-like Mn3CaO4 cluster with a mono-m-oxobridge to a fourth Mn ion and accordingly a mechanism for wateroxidation was proposed (Ferreira et al., 2004). Their study was the

transmembrane a-helices and cofactors in PSII. One monomer of the dimer is showne dotted interface). Chl a head groups and haems are indicated by black wire drawings.helices of D1, D2 and Cyt b-559 are labelled. D1/D2 are highlighted by an ellipse andur prominent landmarks (three irons and the Mn (Mn) cluster) are indicated by arrowsht (2008) by Nature publications.

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first reported structure that included a Ca ion. One year later, B.Loll et al. offered a different structure for the Mn4Ca cluster, thusthe distances between MneMn in the pyramid (three Mn and Ca)were not equal and Mn4 adjoin asymmetrically to the pyramid(Loll et al., 2005). J. Yano et al. considered an extended set ofhypothetical models of the Mn4Ca cluster so that the calculatedEXAFS spectra matched the polarized EXAFS data which wereacquired for PSII crystals (Yano et al., 2006). In 2010, the X-rayEmission Spectroscopy was utilized for the detection of oxo-bridges in the Mn4Ca cluster by analysis of its spectrum(Pushkar et al., 2010). The most considerable information was

Fig. 3. Top the MneCa cluster (circled in red) may be considered as a nanosized MneCa oxidabout w0.5 � 0.25 � 0.25 nm. Adapted from (Umena et al., 2011; Kawakami et al., 2011).

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gained by Shen and Kamiya research groups via the record the PSIIcrystal structure at 1.9 �A resolution (Fig. 3) (Umena et al., 2011;Kawakami et al., 2011). The electron densities illustrated that theWOC cluster is composed of 4 Mn atoms, 1 Ca atom, and 5 oxygenatoms. They showed that 3 Mn and Ca atoms formed four cornersand 4 oxygen atoms occupied the other four corners of a cubane-like structure. The fourth Mn was placed outside of the cubanestructure, and was linked to two Mn atoms within the cubaneby O5. Moreover, four water molecules were bound to theMn4CaO5-cluster (Mn4O5Ca(H2O)4), with two molecules are coor-dinated to Ca and two other to Mn4. Probably, some of these

e in a protein environment Bottom the Mn4O5Ca cluster in PSII that has dimensions of

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molecules are involved in water oxidation. At least 1300 watermolecules were found in a PSII monomer. The formation ofhydrogen bonding by some of these molecules may act as protons,water or oxygen molecule channels. The structural details of theligands around each of Mn and Ca in the cluster are mentioned asfollows. Mn1: three m3-O, two carboxylates (D1-Glu189 andD1-Asp342) and one imidazole group (D1-H332) stabilize theoxidation state of III or IV. Mn2: three m3-O and three bridgingCOO� (D1-Glu342, CP43-Glu 354 and D1-Ala 344) could be sta-bilizer factors of the oxidation number of III or IV. Mn3: three m3-O, one m2-O and two bridging COO� (CP43-Glu 354 and D1-Glu333) groups are present. Four hard m-O ligands could stabi-lize Mn (IV). Mn4: one m4-O, one m2-O, two bridging COO� (D1-Asp170 and D1-Glu333) groups and two H2O molecules are pre-sent. These ligands could stabilize the oxidation state of (III),although deprotonation of water molecules could lead to thestabilization of the oxidation state of (IV). Ca: three m3-O bridges,two bridging COO� (D1-Ala 344 and D1-Asp170) groups and twoH2O molecules are involved.

Recently, crystallographic studies have determined the positionand bonding distances of Ca in the WOC showed a key role of Ca inoxygen evolution (Latimer et al., 1995, 1998; Cinco et al., 1998;Kamiya and Shen, 2003; Ferreira et al., 2004; Lee et al., 2007).The importance of the presence of a Ca ion in theWOCwas revealedin 1984 (Ghanotakis et al., 1984). A. Boussac and A. W. Rutherfordshowed that Ca removal from the cluster hampered the splitting ofwater due to blocking the S2 to S3 transition (Boussac andRutherford, 1988). T. A. Ono and Y. Inoue in 1989 investigated therole of Ca in O2 evolution and found three tasks for it: (i) adjust-ment of oxidation potential (ii) structural regulation of the cluster;(iii) conformational adjustment of the whole water oxidationstructure (Ono and Inoue, 1989). Using XANES, T. A. Ono et al. in1993 considered flash-induced changes the WOC (depleted of Ca).The results indicated that the Mn-cluster and/or its direct ligandcould be oxidized up to two electrons but further events areblocked (Ono et al., 1993).T. A. Ono and T. Noguchi’s groups haveconsidered important issues such as the role of cofactors, structuralchanges for theWOC and so on in PSII by Fourier transform infrareddifference spectroscopy (Noguchi et al., 1992, 1993, 1997, 1999;Noguchi and Sugiura, 2001). In the same year, A. Boussac et al.could deplete the PSII enriched membrane from Ca and found O2evolution did not occur unless Ca ion was added. So, the absence ofCa leads to a disordered Mn cluster (Boussac et al., 1989). Variousstudies have been conducted to find a replacement for Ca and onlySr could restitute the function of water-splitting in Mn4OxSr asMn4OxCa (Ghanotakis et al., 1984; Boussac and Rutherford, 1988;Boussac et al., 2004). This is perhaps due to the similar Lewisacidity of Sr and Ca (Vrettos et al., 2001; Lee and Brudvig, 2004). C.-I. Lee et al. demonstrated a structural role for Ca in the beginning ofS state transitions and also showed that it could be carried out bythe cations with the same ionic radius (Lee et al., 2007). Finally, T.Lohmiller et al. have shown in 2012 that Ca removal does not affectthe electronic properties of the Mn cluster and is not required forstructural preservation of the WOC (Lohmiller et al., 2012). Instead,they proposed that Ca is necessary to preserve the hydrogenbonding between the amino acid tyrosine (Yz) and the Mn cluster.

In addition to direct binding ligands to theWOC, indirect ligandsinclude D1-Asp61, D1-His337, and CP43-Arg357 amino acids thatwere detected in the second coordination domain, in which twoguanidinium h-nitrogen of CP43-Arg357 and the imidazole of D1-His337 are bound to different oxygen groups in the cluster. Moreinformation about the interaction of His337 with the cluster wererevealed by Petrie et al. (Petrie et al., 2012). They found that His337residue interacts with WOC through m3-oxo bridge connectingMn(1), Mn(2) and Mn(3) which may lead to prolongation of Mne

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Mn distances monitored in the PSII crystal structure at 1.9 �A reso-lution. Furthermore, density functional theory displayed thathydrogen bonding of the cluster with His337 residue caused theextension of some of the MneMn distances, specially the distanceof 2.7 �A.

The last known cofactor of the WOC is Cl�. D. I. Arnon and F. R.Whatley in 1949 showed the isolated photosynthetic membranesneed to Cl�, Br�, NO3

� or I� to produce O2 (Arnon andWhatley,1949;Yocum, 2008). Subsequent studies identified Cl� is needed for theelectron donor reactions in the WOC (Bove et al., 1963; Izawa et al.,1969; Yocum, 2008). P. O. Sandusky and C. F. Yocum proposed thatthe chloride ion could act as bridge between Mn atoms (Sanduskyand Yocum, 1984). H. J. Van Gorkom et al. found that supersededions eg. iodide or bromide decreased the stability of the advancedS-states (Wincencjusz et al., 1998). Further experiments revealedCl� was needed to the S2 / S3 and S3 / S4 / S0 transitions of theWOC (Wincencjusz et al., 1999). A role in the regulation of the redoxproperties of WOC was also proposed (Rutherford, 1989). Radio-isotope labeling studies indicated that only one Cl� ion would bebound per each WOC unit (McEvoy and Brudvig, 2006; Lindbergand Andréasson, 1996). K. Olesen and L. -E. Andreasson, usingEPR spectroscopy, suggested that Cl� like Ca could be involved inthe proton transfer away from the WOC (Olesen and Andréasson,2003). The detailed structural data of the WOC obtained by X-raycrystallography at 1.9 �A which was described above, have shownthat two chloride ions are present in the cluster (Umena et al.,2011). The recent mutagenesis studies of D2-K317 by G. W. Brud-vig (Pokhrel et al., 2013) and T. Noguchi (Suzuki et al., 2013) groupshave clearly suggested the possibility that Cl-1 is involved in theproton pathway. I. Rivalta et al. found by structural analysis of theWOC that chloride removal impel the salt-bridge formation be-tween D2-K317 and D1-D61 that may cause the proton transferrepression to the lumen (Rivalta et al., 2011).

3. Mn stabilizing protein auxiliary

The heart of PSII reaction center is consisted of polypeptides D1(PsbA) and D2 (PsbD) which are essentially preserved in theoxygenic photosynthesis species (higher plants, algae, and cyano-bacteria). These proteins are liable for binding of all the redox co-factors involved in electron transfer. Also, PSII includes antennaproteins CP47 (PsbB) and CP43 (PsbC) as well as the two subunits(PsbE and PsbF) of cytochrome b559 (Pagliano et al., 2013). Inaddition, there are several subunits with low molecular mass(10 KDa) which are necessary for stabilizing the assembled PSIIdimer. In contrast to the conservation of the PSII reaction centerand the WOC during the evolution, the subunits stabilizing of theWOC differ between various organisms. The extrinsic proteins ofplants and green algae are PsbO, PsbP and PsbQ. However, cyano-bacteria and diatoms contain PsbO, PsbV and PsbU but red algaeincludes PsbQ in addition to PsbO, PsbV and PsbU (Pagliano et al.,2013; Enami et al., 2008; Bricker et al., 2012). Here, we onlydescribe the PsbA, PsbD, and PsbO proteins of higher plant (formore details of other proteins, see ref (Pagliano et al., 2013)).

3.1. D1 (PsbA)

This protein has a molecular mass of approximate 38 KDa andcontains about 344 amino acids depending on species. PsbA iscomposed of five transmembrane a-helices and two surface a-he-lices (AeE), DE stromal and CD lumenal surface a-helixes (Paglianoet al., 2013; Luci�nski and Jackowski, 2006). PsbAwith PsbD form theheterodimer which is bound to the cofactors involved in PSII-mediated electron transport including: Mn cluster (Asp170,Glu189, His332, Glu333, Asp342, and the C-terminus of Ala344), YZ

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(Tyr161), two pheophytins a (H-bounded by Tyr126 and Glu130),QA, non-heme Fe and QB (interacting via His215 and Ser264) as wellas six chlorophyll a molecules. PD1 is responsible for the chargeseparation and connect to PsbA through His198. Also, excluding thefour mentioned chlorophylls, the PsbA/PsbD heterodimer bindstwo chlorophyll a molecules named ChlZD1 and ChlZD2. Moreover,PsbA binds two chloride ion via D1-Glu333 and D1-Asn338. Asignificant property of the D1 protein is its high susceptibility tophotodamage that leads to decreasing of photosynthetic efficiency.

3.2. D2 (PsbD)

PsbD protein has about 350 amino acids and amolecular mass ofapproximate 39.5 kDa, regarding to the species. It has 5 trans-membrane a-helices (A-E) and two surface a-helices which arelocalized between CeD (luminal) and between D and E (stromal).The D2 protein binds PD2 (by His197), ChlD2 (by a water molecule),the lateral chlorophyll a molecule ChlZD2, QA (His214, Phe261,probably Trp253 and Leu267), and non-heme Fe (interacting viaHis214 and His268). One b-carotene molecule that is possiblyrelevant to PsbD involve in transfer of excitation energy fromChlZD2 to P680. In higher plants, the reversible acetylation andphosphorylation of N-terminal Thr residue can occur for PsbD(Pagliano et al., 2013).

3.3. PsbO

PsbO known as the “Mn-stabilizing’’ protein is present in alloxygen evolving photosynthetic organisms (Miyao and Murata,1984; Najafpour et al., 2012b). It includes 240e257 residuesdepending on species. It has a molecular mass of about 33 kDa. Twomajor domains of PsbO are (i) a filled cylindrical b-barrel structurewhich has a key role in stabilizing the Mn cluster (Ferreira et al.,2004) (ii) a head domain that operates as a docking site. Incontrast to cyanobacteria, PsbO removal from green algae andhigher plant affected the water oxidation efficiency and modulate

Fig. 4. A model for the S state cycle of the Mn cluster of PSII. For clarity, YZ is shown only infor the formation of the S2 state, and their specific functions within the context of the metallwas reprinted with permission from Najafpour et al. (2012b) Copyright (1997) by McMilan

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the Ca and Cle requirements for oxygen evolution (Pagliano et al.,2013; Bricker et al., 2012). The detailed structural survey ofcyanobacterial and plant PSII indicate one and two PsbO per PSIImonomer, respectively. Popelkova et al. showed that plant PsbOhad two sequence motifs at its N-terminus in contrast to cyano-bacterial PsbOwhich had only one sequencemotif (Popelkova et al.,2002; Roose et al., 2007; Nield et al., 2002).

4. Proposed mechanisms for water oxidation in PSII

One of the most important unresolved ambiguity in PSII is thewater oxidation mechanism. Several research groups haveattempted to define an acceptable mechanism by considering thestructural evidence.

The first major studies that led to understand the mechanism ofwater oxidation were done by P. Joliot et al. in 1969. They demon-strated that a four step flash illumination of PSII generates a fluc-tuating pattern of oxygen evolution, in which utmost yield arisesfrom fourth flash. This result was significant since the oxidation oftwo water molecules for the production of one oxygen moleculerequires the removal of four electrons.

Based on this finding, B. Kok et al. in 1970 proposed that in eachof the four stages, a single electron is removed from the WOC(Fig. 1). The accumulated four oxidizing equivalents caused the twowater molecules oxidation (Forbush et al., 1971). Following to theKok cycle, numerous groups represented the proton transferpattern from the WOC into the lumen in the different S-state as:1:0:1:2 (Dau and Haumann, 2008; Pushkar et al., 2008; Suzukiet al., 2009). In 1997, J. Babcock’s group proposed that tyrosine161 may abstract hydrogen atoms from substrate water bound asending ligands to two different Mn ions. In the final stage(hydrogen atom transfer), an OeO bond would be formed betweenthe two terminally coordinated oxides or oxyl radicals (Fig. 4)(Hoganson and Babcock, 1997).

M. Kusunoki in 2007 reviewed the 18O exchange rates of twosubstrate water molecules for S0eS3 states (Kusunoki, 2007). The

the S0 state. Ca and chloride ions are required for oxygen evolution activity, particularlyoradical mechanism are considered in detail elsewhere (Najafpour et al., 2012b). Imagepublications.

anganese oxide in a protein matrix as a natural water-oxidizing site,phy.2014.01.020

Fig. 5. Mono-metallic mechanism. In the mechanism, the OeO bond formation be-tween the oxygens of the two water molecules coordinated to one Mn ion (Popelkovaet al., 2002). Image was reprinted with permission from Popelkova et al. (2002)Copyright (2007) by Elsevier.

Fig. 6. Catalytic cycle of water splitting suggested by DFT QM/MM models of the WOC of PSChanges caused by an S-state transition are highlighted in red. The blue circles highlight submarked in green (Joliot et al., 1969). Image was reprinted with permission from Joliot etreferences to colour in this figure legend, the reader is referred to the web version of this

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formation of 34O2 and 36O2 identified the substrate binding sitesdepending on S-state. He proposed that two substrate water mol-ecules are bound to asymmetric cis-positions on the terminal Mnion being triply bridged (m-oxo, m-carboxylato, and a hydrogenbond) to the Mn cluster (Fig. 5). Also, it was suggested that chlorideion bound to CP43-Arg357 close to Ca ion and D1-His337 may beinvolved in trapping the released proton from the S2-state. Pecoraroet al. suggested the nucleophilic attack of Ca coordinated by hy-droxide to a terminal Mn(V) ¼ O leads to formation of Mn-boundhydroperoxide (Pecoraro et al., 1998).

On the other hand, G.W. Brudvig’s group proposed that the OeOformation is a result of nucleophilic attack of a Ca-bound water onthe electrophilic oxygen atom of a Mn(V) ¼ O species (Brudvig,2008). V. S. Batista et al. using comparison of the DFT QM/MMmodels with high-resolution extended X-ray absorption found thatS3, S4, and S0 states include an additional m-oxo bridge betweenMn(3) and Mn(4) (Fig. 6) (Sproviero et al., 2008). The catalyticreaction is done via an oxyl radical formation by deprotonation ofthe substrate water molecule coordinated to Mn(4). It leads toaccumulation of the fourth oxidizing equivalent in the oxidizedsubstrate water. At last, a nucleophilic attack of substrate watermolecule ligand of Ca to the oxyl radical causes the formation of

II. Dashed arrows indicate transformations leading to the following S state in the cycle.strate water molecules. Coordination bonds elongated by the Jahn-Teller distortion areal. (1969) Copyright (2008) by American Chemical Society. (For interpretation of thearticle.)

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dioxygen. The reaction continues by the replacement of the watermolecules bound to Ca by a water molecule in the second coordi-nation sphere of Ca, deprotonation of the displaced water, releasinga proton to the lumen via CP43-R357, and transferring the otherproton to the basic m-oxo bridge linking Mn(4) and Mn(3).

H. Dau and his colleagues in 2001 discussed the hypotheses ofthe oxidizing transitions as based on EXAFS and XAS data. Theyproposed that during the S2eS3 transition, the mono-m-oxo bridgewas transformed to di-m-oxo bridge along with deprotonation ofhydroxide or water ligand by aid of transition from five-coordinated Mn(III) to six-coordinated Mn(IV) (Fig. 7a) (Dau et al.,2001). Also, EXAFS results revealed that the S0 /S1 transitionincluded deprotonation of a m-oxo-m-hydroxo bridge leading to

Fig. 7. Hypothesis on the S2 / S3 transition. In (A), the Mn complex in its S3-statecontains three unprotonated di-W-oxo bridges; in (B), one of the three di-W-oxobridges is protonated. In (A) and (B), the oxidation of Mn(III) is linked to deprotona-tion of one water-derived ligand (marked by dotted circles) and formation of anadditional bridge between Mn ions. In this scheme, an arrangement of the four Mnions and six bridging ligands has been chosen for the Mn complex in its S2 state; alsoother structural models for the Mn complex in its S2 state models are in agreementwith the EXAFS results (a). Hypothetical scheme on the mechanism of water oxidationand dioxygen formation upon the S3 / S0 transition. Only three Mn ions are depictedbecause the proposed mechanism does not require that four Mn ions are activelyinvolved (Forbush et al., 1971). Optimized transition state for OeO bond formation.

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shortening of MneMn distance from 2.8 to 2.7 �A. Moreover, thechanges in electrochromic absorption showed the charge was notaccumulated by the WOC during the S0 / S1 transition. Theobservation of positive charge accumulation at the WOC could be aresult of charge decompensation by the ligand deprotonation.Recently, P. Glatzel et al. using resonant inelastic X-ray scatteringspectroscopy showed that in the multinuclear clusters like theMn4CaO5 cluster, electrons are strongly delocalized in the cluster,and probably including ligands involved in the redox chemistry(Glatzel et al., 2013).

Per E. M. Siegbahn at Stockholm University used the high-resolution (1.9 �A) structure by Shen et al. (Umena et al., 2011) inhis calculations. The amino acids included in the mode are:

1) The first shell of directly binding amino acids, Asp170, Glu189,His332, Glu333, Asp342, Alal344 and Glu354.

2) The second shell residues Asp61, His337 and Arg357 and thechloride. Lys317 and three water molecules, forming a hydrogenbonding network were also considered in this calculation.

In the proposed mechanism, the S2 / S3 and the S3/S4 tran-sitions start with an electron transfer from YZ to P680þ followed by anexergonic release of a proton of the WOC. Next step in the S2 / S3transition is the oxidation of Mn1.

In this state, the substrate water becomes more strongly boundto Mn1 and the JahneTeller axis on that center disappears. In theS3/S4 transition, there is a proton transfer, which unlike theprevious S-state transition precedes the oxidation step (Fig. 7b).After an oxyl radical formation, there is another proton transferbefore the S4-state state is reached at which OeO bond formationoccurs. In this mechanism, the OeO bond is formed between anoxyl radical in the center of the cluster and a Mn-bridging m-oxoligand (Fig. 7b) (Suzuki et al., 2009).

5. Artificial photosynthesis based on synthetic catalysts

As mentioned before, the researchers have been exploiting theobtained structural details of PSII to reach an efficient catalystcompound for water oxidation and as a result hydrogen production,an appropriate alternative to fossil fuels. To this end, the main focusis on the WOC (Najafpour, 2011; Najafpour et al., 2011; Hou et al.,2010).

There are some reports for PSII extraction and its immobiliza-tion on the surface of electrodes (Ataka et al., 2004; Badura et al.,2008, 2006; Kato et al., 2012; Kato et al., 2013). These PSII coatedelectrodes act as a heterogeneous catalyst for water oxidation(Fig. 8) (Badura et al., 2011). Under illumination, they have theability for water splitting at oxidation potentials as low as w0 V vs.NHE (Badura et al., 2008). However, the low operational stability ofPSII under light exposure limits their use. Therefore, this is themainreason to synthesize new catalysts instead of the direct use of PSII.

There are some aspects that should be considered for synthesisof new catalysts: i) facilitation of electron transfer between the

Fig. 8. Basic principle for the integration of PSII into bioelectrochemical devices(Brudvig, 2008).

anganese oxide in a protein matrix as a natural water-oxidizing site,phy.2014.01.020

Fc

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catalyst and an oxidizing equivalent that could be done bydecreasing the oxidation potentials of the catalyst. ii) balancingbetween reducing the redox potential of the catalyst and the sus-tainability of oxidizing power for water oxidation. iii) non-toxicityand cheapness. Among the introduced catalysts, the preferredones contain Fe, Ni, Ru, Co, or Mn transition metals (Xu et al., 2011;Du and Eisenberg, 2012; Soriano-López et al., 2013; Najafpour et al.,2012c).

Based on recent high resolution crystallography of PSII, it isinteresting that despite the existence of hundreds amino acidsaround the Mn cluster, only 3-4 residue are directly coordinated tothe cluster. The possible roles of these residues are regulation ofcharges and stabilizing the cluster. Besides, the activity of the WOCis decreased in the absence of specific amino acids (Debus, 2008).Thus, the synthetic super anode compounds should use strategicgroups for facilitation of proton transfer, regulation of charges, andhelp in coordination of water molecule at appropriate sites(Najafpour et al., 2012a).

In catalyst optimization, measurements are usually done bydriving the WOC with a sacrificial oxidant. The oxidants likecerium(IV), ruthenium(III) tris(bipyridine) cation, sodium perox-odisulfate, potassium peroxymonosulfate, and sodium periodateallow the production of abundant oxygen in bulk solution.

Recently, G. W. Brudvig’s group published a tutorial review andextensively surveyed the advantages and disadvantages of different

ig. 9. Light induced water splitting by PSII in photosynthesis and hydrogen productionomplexes and a possible scheme for mimicking the natural process. (Badura et al., 2011).

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oxidants (Parent et al., 2013). Also, electrochemical methods areuseful to activate the catalyst by applying potential and probe thewater oxidation (Dinc�a et al., 2010; Parent et al., 2013; Bediakoet al., 2013; Bergmann et al., 2013). Other method for productionof H2 is light-drivenwater oxidation at PSII which can be connectedto H2 production by hydrogenase (an enzyme in the many greenalgae and cyanobacteria that can convert the released protons intodihydrogen). A general plan for H2 production usingwater and solarenergy is shown in Fig. 9 (Lubitz et al., 2008), inwhich two dyes area representative of the two photosystems in oxygenic photosyn-thesis. At first, a dye molecule (A) that mimics PSII absorbs thevisible wavelength of light and then the photoexcited dye (A*)reduces a redox mediator (R). The hole produced in A is transferredto the adjacent water-oxidation catalyst (Catox).

In the PSI mimic, a photoexcited dye (D*) reduces the proton-reducing catalyst. The reduced redox mediator provides electronsto the oxidized dye like plastocyanin in the natural PSI. In this way,light-driven water oxidation is coupled with light-driven hydrogenproduction (Lubitz et al., 2008; Pokhrel and Brudvig, 2013). Somegroups have reported different Mn oxides as catalysts toward wateroxidation (Morita et al., 1977; Shafirovich and Shilov, 1979;Harriman et al., 1988; Najafpour et al., 2012c). In 2010, a newMneCa oxide catalyst by oxidation of Mn(II) ions in the presence ofKMnO4 was synthesized which was inspired by the MneCa clusterstructure in PSII (Najafpour et al., 2010). The next results showed

by an [FeFe] hydrogenase shown together with the structure of respective protein

anganese oxide in a protein matrix as a natural water-oxidizing site,phy.2014.01.020

Fig. 12. Cyclic voltammograms (CVs) of a PVPePt electrode (magenta), l-MnOxePVPePt (red), m-MnOxePVPePt (yellow), h-MnOxePVPePt (grey), and MnOx on Pt (blue)(LiClO4 in water (0.1 M), pH ¼ 6.3) at a scan rate of 100 mV s�1 in both 0.0e2.0 V and1.0e1.4 V. The range of Mn(III) to Mn(IV) oxidation is 0.6e0.9 V. The broadening ofthese redox transitions is most probably due to electronic interactions of Mn sites inthe material. The black arrow indicates the potential at which the oxygen measure-ments were performed (Najafpour et al., 2010). Image was reprinted with permissionfrom Najafpour et al. (2010) Copyright (2013) by RSC publication. (For interpretation ofthe references to colour in this figure legend, the reader is referred to the web versionof this article.)

Fig. 10. TEM (a) and HRTEM (b) of nanolayered MneCa oxide.

M.M. Najafpour et al. / Plant Physiology and Biochemistry xxx (2014) 1e1310

that nanolayeredMn oxides (Fig.10) are efficient catalysts for wateroxidation (Najafpour and Allakhverdiev, 2012; Najafpour et al.,2012d, 2013a,b; Birkner et al., 2013; Najafpour and Sedigh, 2013).However, these model systems contain no groups analogous toamino acids, as exist in PSII. There are a few reports of the synthesis

Fig. 11. TEM image of l-MnOxePVP (Najafpour et al., 2010) Image was reprinted withpermission from Najafpour et al. (2010) Copyright (2013) by RSC publication.

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of new Mn oxides-based catalysts inside an organic compoundmatrix as a proper alternative for the amino acids (Najafpour et al.,2012e, 2013c,d,e; Najafpour and Moghaddam, 2012; Takashimaet al., 2012; Singh et al., 2013).

Recently, Najafpour et al. introduced a novel efficient nano-layered Mn oxide/poly(4-vinylpyridine) catalyst with only 50 mVoverpotential needed for water oxidation (Najafpour et al., 2013e).The reasons of using poly(4-vinylpyridine) (PVP) were: i) providinga suitable buffered medium for Mn oxide. ii) the operation of pyr-idine groups in the polymer as proton acceptor, Mn (III) stabilizer,and also overpotential reducer for water. iii) the polymer stability

Fig. 13. Oxygen evolution by l-MnOx-PVP-Pt at 700 mV (vs. Ag/AgCl) with only 50 mVoverpotential. Oxygen evolution by l-MnOx-PVP-Pt at potential of 700 mV using pure(red) and Persian Gulf (black). For water from Persian Gulf, no LiClO4 was added. (Forinterpretation of the references to colour in this figure legend, the reader is referred tothe web version of this article.)

anganese oxide in a protein matrix as a natural water-oxidizing site,phy.2014.01.020

M.M. Najafpour et al. / Plant Physiology and Biochemistry xxx (2014) 1e13 11

against strong oxidants. iv) the conductivity and polyelectrolyteproperties of the polymer. Various compounds with different Mnoxide and a certain PVP values (l-MnOxePVP, m-MnOxePVP, h-MnOxePVP) were synthesized. TEM images illustrated the layeredstructure of Mn oxides which is the optimal structure for artificialphotosystem (Fig.11). As illustrated in Fig.12, similar to natural PSII,the one with low MnOx and high PVP contents show lower over-potential towardwater oxidation on the platinum electrode surfaceat near natural pH. Moreover, oxygen evolution occurred at 700 mV(Fig. 13). Also, they found that the mentioned catalyst could applysuccessfully for the Persian Gulf water oxidation with a turnoverfrequency of 0.07 s�1. This is a great outcome because in thepresence of catalyst, the harmful chlorine gas was not formed onthe anode. So, such catalysts open the way for H2 fuel productionfrom the great source of water oceans in the future.

Artificial solar fuel apply the basic principles of photosynthesis,such as light harvesting, charge separation between donor andacceptor molecules, electron transfer, and the multi-electron cat-alytic processes to design systems for cheap and environmentallyfriendly fuel production by renewable energy sources. Among ofdifferent reactions, water splitting is very promising (Alibabaeiet al., 2013; Kern et al., 2013; Nocera, 2012).

6. Conclusion

Synthesis of new catalysts for the oxidation of water usingenvironmentally friendly materials is a big challenge in order toachieve clean fuel. Artificial solar fuel applying the basic principlesof photosynthesis to design systems for fuel production byrenewable energy sources. The art of water oxidation by PSII can beconsidered in artificial photosynthesis. This purpose is accom-plished by a precise look at the Mn cluster structure found in PSII.Mn oxides are promising compounds for water oxidationwith highefficiency.

Acknowledgments

Authors (MMN, MZG, BH) are grateful to Institute for AdvancedStudies in Basic Sciences and the National Elite Foundation forfinancial support. This work was also supported by grants from theRussian Foundation for Basic Research (Nos: 13-04-91372; 14-04-01549; 14-04-92690), and by Molecular and Cell Biology Programsof the Russian Academy of Sciences, and by BMBF (no. 8125)Bilateral Cooperation between Germany and Russia to SIA. Thiswork was also supported by Grant-in-Aids for Scientific Researchfrom the Ministry of Education of Japan (21570038 and 22370017),and grant from JST PRESTO to TT.

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