Structural rearrangements preceding dioxygen formation by the water oxidation complex of photosystem II

Photosynthetic water oxidation is catalyzed by the Mn₄CaO₅ cluster of photosystem II. Recent studies implicate an oxo bridge atom, O5, of the Mn₄CaO₅ cluster, as the “slowly exchanging” substrate water molecule. The D1-V185N mutant is in close vicinity of O5 and known to extend the lag phase and retard the O₂ release phase (slow phase) in this critical last S3+→S0 transition of water oxidation. The pH dependence, hydrogen/deuterium (H/D) isotope effect, and temperature dependence on the O₂ release kinetics for this mutant were studied using time-resolved O₂ polarography, and comparisons were made with WT and two mutants of the putative proton gate D1-D61. Both kinetic phases in V185N are independent of pH and buffer concentration and have weaker H/D kinetic isotope effects. Each phase is characterized by a parallel or even lower activation enthalpy but a less favorable activation entropy than the WT. The results indicate new rate-determining steps for both phases. It is concluded that the lag does not represent inhibition of proton release but rather, slowing of a previously unrecognized kinetic phase involving a structural rearrangement or tautomerism of the S₃⁺ ground state as it approaches a configuration conducive to dioxygen formation. The parallel impacts on both the lag and O₂ formation phases suggest a common origin for the defects surmised to be perturbations of the H-bond network and the water cluster adjacent to O5.Oxygenic photosynthesis depends on the light-driven water-plastoquinone oxidoreductase, photosystem II (PSII), which catalyzes the complete oxidation of H₂O molecules to O₂. The electrons extracted from H₂O oxidation are the reductant for autotrophic metabolism, and the liberated protons contribute to the transmembrane proton motive force powering ATP production (reviewed in refs. 1–4). The byproduct, O₂, has transformed our Earth’s atmosphere and makes heterotrophic life in the biosphere possible. A metal cluster, the Mn₄CaO₅, functions as the catalytic site of H₂O oxidation and is buried in a protein domain of PSII referred to as the H₂O oxidation complex (WOC). The primary photochemical electron donor of the reaction center (RC) complex is a dimeric chlorophyll moiety designated P₆₈₀ that is coordinated by the pseudosymmetrically arranged D1 and D2 polypeptides. Together, the D1/D2 heterodimer are responsible for the coordination of most of the photochemically active cofactors of the RC. Photoexcitation of P₆₈₀ initiates transmembrane charge separation with the activated electron moving to plastoquinone species, Q_A and Q_B, on the acceptor side of the RC. The resultant electron hole (P680+•) functions as the powerful oxidant for the extraction of electrons from substrate H₂O bound to the Mn₄CaO₅ (Fig. 1A). However, the oxidation of the Mn₄CaO₅ occurs through a redox active tyrosyl of the D1 protein, D1-Tyr161 (Y_Z), located between P₆₈₀ and the metal cluster. Thus, Y_Z functions as a secondary donor to the photochemical RC and the primary oxidant of the Mn₄CaO₅. Oxidation of Y_Z creates a neutral tyrosine radical YZ• because of a coupled proton transfer from the tyrosine to the nearby D1-His190, which is H-bonded to Y_Z. Accordingly, Y_Z oxidation leads to the formation of the positive charged pair YZ• His190⁺ by moving the proton within the H bond (5–9). According to the “proton rocking” mechanism, this proton returns to Y_Z on its reduction by the Mn₄CaO₅ cluster. The transient formation of the positive charge in the immediate vicinity of the cluster may have a critical electrostatic effect in promoting proton transfer and/or H-bond rearrangements elsewhere in the WOC.Open in a separate windowFig. 1.(A) Mutations constructed in Synechocystis sp. PCC6803 to D1 residue Val185 (rendered with yellow sticks) and Val185Asn substitution showing one possible configuration of the amide extending into the water cavity H bonding (red dashes) with the water molecules (cyan spheres) close to oxo bridge atom, O5 (bright blue), of the Mn₄CaO₅ cluster. Rudimentary analysis of crystal structure with PyMol indicates multiple rotamer configurations of the asparagine side chain without steric clashes, except displacements of the water molecules. The amide moiety introduces four potential H-bonding sites: two acceptor (-C = O) and two donor (-NH₂) sites. Other energetically feasible rotamers may interact with the essential chloride anion cofactor (Cl1; green). (B) A large retardation of O₂ release kinetics during S3+→S0n was found in D1-V185N mutant (23), similar to D1-D61N and D1-D61A mutants (16, 20, 32). (C) Proposed intermediates after excitation of PSII in the S3+YZ• state. Step 1: oxidation of Y_Z by P₆₈₀₊ forming S3+YZ•. Step 2: loss of the proton forming S3nYZ•. Step 3: reduction of YZ•/oxidation of the Mn cluster. Step 4: reduction of the cluster by oxidation of substrate. Step 5: O-O bond formation. Step 6: H₂O binding and O₂ release. The S₄ state. (D) Representation of the entire S-state cycle, emphasizing the alternating electron and proton removal from the cluster. Classic Kok S-states correspond to those shown in red outline.Charge separation in the photochemical RC is a single-electron process, but it must be coupled with the four-electron oxidation: 2H₂O→O₂ + 4e⁻ + 4H⁺. PSII solves the valence mismatch problem by accumulating oxidizing equivalents within the Mn₄CaO₅ cluster as deduced from early observations of period 4 oscillatory O₂ release under flash illumination (10) that led to the formulation of a model invoking storage (S) states consisting as four semistable states produced by flash illumination (S₀, S₁, S₂, and S₃) and a transient state (S₄) (11). O₂ formation and release occurs during the S₃ → S₀ transition, with a hypothetical intermediate S₄ that forms on oxidation of S₃ followed by the formation of O₂ and the reduction of Mn₄CaO₅ by substrate H₂O (11). Proton release to the lumen of the thylakoids occurs at specific points during the catalytic cycle (reviewed in ref. 12). Deprotonation minimizes the accumulation of positive charge on the Mn₄CaO₅ cluster and provides a “redox leveling” effect that ensures that successive withdrawals of electrons from the metal center remain energetically feasible through all of the S states of the catalytic cycle. Detailed analyses have revealed an alternation of electron transfer and proton transfer events (13, 14), leading to the reformulation for the cycle, where the indices n and + refer to the electrostatically neutral and positive states, respectively, relative to the S₁ⁿ state (Fig. 1D). Because of the paucity of identified reaction intermediates in the S3+→S0n transition, the mechanism of water oxidation by PSII still remains elusive. In this step, the O-O bond is formed, O₂ and two protons are released, and at one substrate, water binds to the Mn₄CaO₅ cluster. Possible intermediates in this poorly understood final stage are depicted in Fig. 1C. A 30- to 250-μs lag phase preceding the release of O₂ and the reduction of the Mn₄CaO₅ has been assigned to the release of a proton (step 2 in Fig. 1C) (5, 13–19). Step 3 in Fig. 1C refers to the oxidation of the Mn cluster by YZ• and has been observed to initiate 250 μs after the excitation of the S₃⁺ ground state (18). The initiation of step 3 marks the end of the lag phase and leads to formation of the transient and chemically undefined S₄ state. It should be pointed out that the lag phase is multiphasic (13), and thus, step 2 and perhaps, step 3 in Fig. 1C consist of more than one process—a point that is significant to the experiments described below. Mutational analysis (16, 20–23) and substitution of the Ca²⁺ and Cl⁻ cofactors (24) result in a slowdowns of the S3+→S0n transition, including an increase in the duration of the lag phase. These slowdowns in the lag phase could be attributed to an impairment of proton release to the solvent bulk, but as shown in this work, they may be better ascribed to a slowdown of some other molecular rearrangement that precedes the release of O₂ and becomes rate-determining in the mutants studied here.As shown in Fig. 1A, the Mn₄CaO₅ cluster resembles a distorted chair, where four oxygen atoms link the three Mn atoms and one Ca atom together by μ-oxo bridges to form the cuboidal unit. The fourth dangling Mn (Mn4) is located outside the cube and linked to one of the Mn corners (Mn3) by a μ-oxo bridge ligation (O4) and a fifth bridging μ-oxo (O5) to the Ca (25, 26). The bonding arrangement of O5 is proposed to be flexible and consequently, provides flexibility to the entire structure (27): The S₂⁺ state may exist in two nearly isoenergetic configurations as suggested by the alternative EPR signals (27): a low-spin open cube, where O5 is bonded to Mn4 and not Mn1 as depicted in Fig. 1A, or the high-spin, closed cube state, where O5 is bonded to Mn1 and not Mn4. The slowly exchanging substrate water is assigned to O5, and the envisioned structural flexibility would account for the unexpectedly fast rate of exchange of O5 with external water (28). The Mn₄CaO₅ cluster is ligated to the D1 and CP43 subunits of PSII by six carboxylate ligands and one histidine. Other than the first sphere ligands, the geometry of the Mn₄CaO₅ cluster is also maintained by second sphere amino acids and a number of water molecules situated in the cavity of the catalytic site. Water molecules not only are substrate but also, participate in a reticulated H-bond network (HBN), interacting the first and second sphere ligands of the Mn₄CaO₅ cluster and forming paths connecting the cluster with the external aqueous phase of the thylakoid lumen (reviewed in ref. 29). This HBN includes the participation of second sphere ligand D1-Asp61, which may be the initial residue of a main proton egress pathway through its interaction with a water molecule designated W1 (30). Recently, point mutations were generated to interfere with the water molecules surrounding the Mn₄CaO₅ cluster by altering the shape and polarity of the protein surface facing the aqueous pockets (23). Mutations of a valine, D1-V185, exhibit the most interesting phenotypes. D1-Val185 is part of the broad channel, because it passes the Mn₄CaO₅ cluster and is in the close vicinity of O5 (31) (Fig. 1A). The D1-V185N mutation has a strong effect on the oxygen evolution. In contrast to the short lag (<250 µs) and short half-lifetime of 1−2 ms of the S3+→S0n transition in the WT, the mutant has a drastically prolonged lag phase followed by a drastically slowed rate of oxygen release (Fig. 1B; see below), which opens a new window for the study of the reaction pathway during the dioxygen-forming stage of the catalytic cycle. Here, we carefully characterized the pH and temperature dependence and deuterium isotope effect on the O₂ release kinetics of the D1-V185N mutant and made comparison with the WT and D1-D61N and D1-D61A mutants (16, 20, 32). The goal is to gain insight into the nature of the elusive intermediates formed during the lag and O₂ release phases of the S3+→S0n transition.     

Improving the efficiency of water splitting in dye-sensitized solar cells by using a biomimetic electron transfer mediator

Photoelectrochemical water splitting directly converts solar energy to chemical energy stored in hydrogen, a high energy density fuel. Although water splitting using semiconductor photoelectrodes has been studied for more than 40 years, it has only recently been demonstrated using dye-sensitized electrodes. The quantum yield for water splitting in these dye-based systems has, so far, been very low because the charge recombination reaction is faster than the catalytic four-electron oxidation of water to oxygen. We show here that the quantum yield is more than doubled by incorporating an electron transfer mediator that is mimetic of the tyrosine-histidine mediator in Photosystem II. The mediator molecule is covalently bound to the water oxidation catalyst, a colloidal iridium oxide particle, and is coadsorbed onto a porous titanium dioxide electrode with a Ruthenium polypyridyl sensitizer. As in the natural photosynthetic system, this molecule mediates electron transfer between a relatively slow metal oxide catalyst that oxidizes water on the millisecond timescale and a dye molecule that is oxidized in a fast light-induced electron transfer reaction. The presence of the mediator molecule in the system results in photoelectrochemical water splitting with an internal quantum efficiency of approximately 2.3% using blue light.     

Photochemical oxidation of water and reduction of polyoxometalate anions at interfaces of water with ionic liquids or diethylether

Photoreduction of [P(2)W(18)O(62)](6-), [S(2)Mo(18)O(62)](4-), and [S(2)W(18)O(62)](4-) polyoxometalate anions (POMs) and oxidation of water occurs when water-ionic liquid and water-diethylether interfaces are irradiated with white light (275-750 nm) or sunlight. The ionic liquids (ILs) employed were aprotic ([Bmim]X; Bmim = (1-butyl-3-methylimidazolium, X = BF(4), PF(6)) and protic (DEAS = diethanolamine hydrogen sulphate; DEAP = diethanolamine hydrogen phosphate). Photochemical formation of reduced POMs at both thermodynamically stable and unstable water-IL interfaces led to their initial diffusion into the aqueous phase and subsequent extraction into the IL phase. The mass transport was monitored visually by color change and by steady-state voltammetry at microelectrodes placed near the interface and in the bulk solution phases. However, no diffusion into the organic phase was observed when [P(2)W(18)O(62)](6-) was photo-reduced at the water-diethylether interface. In all cases, water acted as the electron donor to give the overall process: 4POM + 2H(2)O + hν → 4POM(-) + 4H(+) + O(2). However, more highly reduced POM species are likely to be generated as intermediates. The rate of diffusion of photo-generated POM(-) was dependent on the initial concentration of oxidized POM and the viscosity of the IL (or mixed phase system produced in cases in which the interface is thermodynamically unstable). In the water-DEAS system, the evolution of dioxygen was monitored in situ in the aqueous phase by using a Clark-type oxygen sensor. Differences in the structures of bulk and interfacial water are implicated in the activation of water. An analogous series of reactions occurred upon irradiation of solid POM salts in the presence of water vapor.     

Stable solar-driven oxidation of water by semiconducting photoanodes protected by transparent catalytic nickel oxide films

Reactively sputtered nickel oxide (NiO_x) films provide transparent, antireflective, electrically conductive, chemically stable coatings that also are highly active electrocatalysts for the oxidation of water to O₂(g). These NiO_x coatings provide protective layers on a variety of technologically important semiconducting photoanodes, including textured crystalline Si passivated by amorphous silicon, crystalline n-type cadmium telluride, and hydrogenated amorphous silicon. Under anodic operation in 1.0 M aqueous potassium hydroxide (pH 14) in the presence of simulated sunlight, the NiO_x films stabilized all of these self-passivating, high-efficiency semiconducting photoelectrodes for >100 h of sustained, quantitative solar-driven oxidation of water to O₂(g).The oxidation of water to O₂(g) is a critical process for the sustainable solar-driven generation of fuels, including the generation of carbon-based fuels by the solar-driven reduction of carbon dioxide as well as the generation of H₂(g) by solar-driven water splitting (1, 2). Many technologically important semiconductors, including silicon (Si), Group III–V materials such as gallium arsenide (GaAs), and Group II–VI materials such as cadmium telluride (CdTe), have optimal band gaps for use in an integrated, dual light-absorber, solar-fuels generator (3). However, these materials are generally unstable and corrode or passivate rapidly when operated under photoanodic conditions in aqueous electrolytes. Furthermore, the efficient operation of a passive and intrinsically safe water-splitting system requires the use of either strongly alkaline or acidic electrolytes, presenting additional constraints on the stability of the photoanodes and electrocatalysts (4–6).The search for new, stable compound semiconductors or molecular systems for water oxidation has thus far yielded materials with low efficiencies and/or limited stability (7, 8). An alternative strategy for addressing the lack of materials known to be stable under the conditions needed for the efficient oxidation of water is to protect high-efficiency, technologically important semiconductors to enable their use in integrated solar-fuels generators by using buried semiconductor junctions to form a photovoltaic (PV)-biased electrosynthetic cell (9). In this approach, the surface of the otherwise unstable semiconducting light absorber is covered by a layer of a stable and partially transparent conductive oxide or metal, which serves either as a Schottky barrier or as a transparent conductive contact to a photoelectrode that contains a buried junction to provide the requisite charge separation. Metallized contacts to radial p-n junctions in Si microspheres have been used to effect the unassisted solar-driven splitting of HI(aq) and HBr(aq) (10), and metallized contacts have been used in conjunction with triple-junction–based hydrogenated amorphous Si (a-Si:H) photovoltaics for PV-biased electrosynthetic water splitting (11–13). The protective layers used in an integrated photoanode generally require a separate electrocatalyst for the oxidation of water on the electrode surface. Further, the entire assembly must be chemically compatible with and stable in the electrolytes and at the electrode potentials associated with photoelectrochemical water oxidation (14). Metals, metal alloys, semiconductors, degenerately doped transparent conducting oxides, catalytic transition-metal compounds, organic polymers, and surface functionalization methods have all been explored for this purpose, with only limited stability, limited electrical properties at junctions, and/or limited activity for water oxidation observed to date (14).Sputtered nickel oxide (NiO_x) films have been recently shown to be optically transparent, antireflective, conductive, stable, and highly catalytically active while protecting n-Si and np⁺-Si photoanodes in contact with aqueous 1.0-M KOH for the photoelectrochemical oxidation of water (15, 16). For np⁺-Si photoanodes, such an approach allowed for the stable production of O₂(g) for >1,200 h of continuous operation under 1-Sun simulated solar illumination, with a photocurrent-onset potential of −180 mV relative to the equilibrium water-oxidation potential and current densities in excess of 29 mA⋅cm⁻² at the equilibrium water-oxidation potential. We demonstrate herein that this protection strategy can be extended to semiconducting materials used in commercial photovoltaic solar cells and in high-efficiency devices, specifically textured crystalline Si passivated with a layer of amorphous Si, known as a heterojunction Si (HTJ-Si) device with a typical structure of (p⁺-a-Si|i-a-Si|n-c-Si|i-a-Si|n⁺-a-Si); n-i hydrogenated amorphous Si (a-Si:H) structures; and single-crystalline n-type CdTe light absorbers. All of these materials have band gaps appropriate (1–2 eV) for use in integrated solar fuel-generation devices, and this work demonstrates that the NiO_x protection strategy is effective on both single-crystalline and noncrystalline semiconductors.     

Spectroelectrochemical determination of the redox potential of pheophytin a,the primary electron acceptor in photosystem II

Thin-layer cell spectroelectrochemistry, featuring rigorous potential control and rapid redox equilibration within the cell, was used to measure the redox potential E_m(Phe a/Phe a⁻) of pheophytin (Phe) a, the primary electron acceptor in an oxygen-evolving photosystem (PS) II core complex from a thermophilic cyanobacterium Thermosynechococcus elongatus. Interferences from dissolved O₂ and water reductions were minimized by airtight sealing of the sample cell added with dithionite and mercury plating on the gold minigrid working electrode surface, respectively. The result obtained at a physiological pH of 6.5 was E_m(Phe a/Phe a⁻) = −505 ± 6 mV vs. SHE, which is by ≈100 mV more positive than the values measured ≈30 years ago at nonphysiological pH and widely accepted thereafter in the field of photosynthesis research. Using the P680* − Phe a free energy difference, as estimated from kinetic analyses by previous authors, the present result would locate the E_m(P680/P680⁺) value, which is one of the key parameters but still resists direct measurements, at approximately +1,210 mV. In view of these pieces of information, a renewed diagram is proposed for the energetics in PS II.     

Structural changes in the Mn4Ca cluster and the mechanism of photosynthetic water splitting

Photosynthetic water oxidation, where water is oxidized to dioxygen, is a fundamental chemical reaction that sustains the biosphere. This reaction is catalyzed by a Mn₄Ca complex in the photosystem II (PS II) oxygen-evolving complex (OEC): a multiprotein assembly embedded in the thylakoid membranes of green plants, cyanobacteria, and algae. The mechanism of photosynthetic water oxidation by the Mn₄Ca cluster in photosystem II is the subject of much debate, although lacking structural characterization of the catalytic intermediates. Biosynthetically exchanged Ca/Sr-PS II preparations and x-ray spectroscopy, including extended x-ray absorption fine structure (EXAFS), allowed us to monitor Mn–Mn and Ca(Sr)–Mn distances in the four intermediate S states, S₀ through S₃, of the catalytic cycle that couples the one-electron photochemistry occurring at the PS II reaction center with the four-electron water-oxidation chemistry taking place at the Mn₄Ca(Sr) cluster. We have detected significant changes in the structure of the complex, especially in the Mn–Mn and Ca(Sr)–Mn distances, on the S₂-to-S₃ and S₃-to-S₀ transitions. These results implicate the involvement of at least one common bridging oxygen atom between the Mn–Mn and Mn–Ca(Sr) atoms in the O–O bond formation. Because PS II cannot advance beyond the S₂ state in preparations that lack Ca(Sr), these results show that Ca(Sr) is one of the critical components in the mechanism of the enzyme. The results also show that Ca is not just a spectator atom involved in providing a structural framework, but is actively involved in the mechanism of water oxidation and represents a rare example of a catalytically active Ca cofactor.     

Efficient water oxidation catalyzed by homogeneous cationic cobalt porphyrins with critical roles for the buffer base

A series of cationic cobalt porphyrins was found to catalyze electrochemical water oxidation to O₂ efficiently at room temperature in neutral aqueous solution. Co–5,10,15,20-tetrakis-(1,3-dimethylimidazolium-2-yl)porphyrin, with a highly electron-deficient meso-dimethylimidazolium porphyrin, was the most effective catalyst. The O₂ formation rate was 170 nmol⋅cm⁻²⋅min⁻¹ (k_obs = 1.4 × 10³ s⁻¹) with a Faradaic efficiency near 90%. Mechanistic investigations indicate the generation of a Co^IV-O porphyrin cation radical as the reactive oxidant, which has accumulated two oxidizing equivalents above the Co^III resting state of the catalyst. The buffer base in solution was shown to play several critical roles during the catalysis by facilitating both redox-coupled proton transfer processes leading to the reactive oxidant and subsequent O–O bond formation. More basic buffer anions led to lower catalytic onset potentials, extending below 1 V. This homogeneous cobalt-porphyrin system was shown to be robust under active catalytic conditions, showing negligible decomposition over hours of operation. Added EDTA or ion exchange resin caused no catalyst poisoning, indicating that cobalt ions were not released from the porphyrin macrocycle during catalysis. Likewise, surface analysis by energy dispersive X-ray spectroscopy of the working electrodes showed no deposition of heterogeneous cobalt films. Taken together, the results indicate that Co–5,10,15,20-tetrakis-(1,3-dimethylimidazolium-2-yl)porphyrin is an efficient, homogeneous, single-site water oxidation catalyst.Water oxidation is a key step in photosynthesis that efficiently harvests and stores solar energy (1). The oxidation of H₂O to O₂ is a four-electron, four-proton process in which O–O bond formation is the key chemical step. In photosystem II, these proton-coupled electron transfers (PCETs) occur via a tyrosine at the Mn₄Ca oxygen-evolving complex (2). An important thermodynamic aspect of photosynthesis is the efficient conversion of photonic energy to electrical potential, thus providing 99% of the driving force required to convert CO₂ to carbohydrates (Eqs. 1 and 2):andThe development of synthetic catalysts that can mediate water oxidation under mild conditions with a minimal energy cost has become an appealing challenge for chemists (3–7). Among various approaches, homogeneous molecular catalysts have shown attractive features such as controllable redox properties, ease of investigating reaction mechanisms, and strategies for the characterization of reactive intermediates (8, 9). Recently, such efforts have resulted in the development of a significant number of systems based on single-site and multinuclear transition metal complexes including Mn, Fe, Co, Cu, Ru, and Ir (9–15). Examples of cobalt-based molecular catalysts include a cobalt phthalocyanine (16), a cobalt “hangman” corrole (17), multipyridine cobalt complexes (18, 19), a dinuclear Co-peroxo species (20), and most recently, Co-porphyrins (21). Determining if these molecular complexes retain their homogenous nature during catalysis or merely act as precursors of truly active heterogeneous species such as films and nanoparticles has proven to be problematic (8, 22, 23). This scenario is especially critical for Co-based molecular catalysts because heterogeneous CoO_x species are highly active, so that even small amounts of this surface oxide layer could contribute significantly to the catalytic activity (24–27). Further, very little detailed mechanistic information for these systems is available to date, and the assignment of the key oxidants responsible for oxidizing a water molecule has been challenging (28). Here we describe mechanistic studies of a series of water-soluble, cationic cobalt porphyrins that effectively catalyze water oxidation electrochemically under mild conditions at neutral pH. The homogeneity of these catalysts was confirmed by a variety of techniques including electrochemical, spectroscopic, and surface analysis that exclude the alternative formation of heterogeneous cobalt oxide films. Our results indicate the generation of a reactive, high-valent Co^IV-porphyrin cation radical that has accumulated two oxidizing equivalents above the resting Co^III catalyst. Further, we demonstrate a critical role for the buffer base that accepts a proton during the O–O bond-formation event.     

Electron-transfer sensitization of H2 oxidation and CO2 reduction catalysts using a single chromophore

Energy-storing artificial-photosynthetic systems for CO₂ reduction must derive the reducing equivalents from a renewable source rather than from sacrificial donors. To this end, a homogeneous, integrated chromophore/two-catalyst system is described that is thermodynamically capable of photochemically driving the energy-storing reverse water–gas shift reaction (CO₂ + H₂ → CO + H₂O), where the reducing equivalents are provided by renewable H₂. The system consists of the chromophore zinc tetraphenylporphyrin (ZnTPP), H₂ oxidation catalysts of the form [Cp^RCr(CO)₃]^–, and CO₂ reduction catalysts of the type Re(bpy-4,4′-R₂)(CO)₃Cl. Using time-resolved spectroscopic methods, a comprehensive mechanistic and kinetic picture of the photoinitiated reactions of mixtures of these compounds has been developed. It has been found that absorption of a single photon by broadly absorbing ZnTPP sensitizes intercatalyst electron transfer to produce the substrate-active forms of each. The initial photochemical step is the heretofore unobserved reductive quenching of the low-energy T₁ state of ZnTPP. Under the experimental conditions, the catalytically competent state decays with a second-order half-life of ∼15 μs, which is of the right magnitude for substrate trapping of sensitized catalyst intermediates.The inexorable growth of global energy consumption and concern over its attendant environmental consequences has spurred considerable research into developing means to store solar energy in the form of renewable chemical fuels (1, 2). Among the potential feedstocks for these solar fuels, CO₂ is a desirable target because it is the end product of the combustion of fossil fuels. Thus, developing solar-driven mechanisms for chemically reducing CO₂ to energy-rich products holds the potential to recycle conventional fuels and mitigate their carbon impact (3–7).Numerous studies over the past 30 y have investigated homogeneous artificial-photosynthetic systems for CO₂ reduction, in which a photoexcited chromophore accomplishes the transfer of electrons from a source of reducing equivalents to a CO₂ reduction catalyst (8–14). With very rare exceptions (15), the reducing equivalents consumed in these photochemical CO₂ reduction reactions have been supplied by sacrificial electron donors. These reagents are used because their prompt decomposition following photoinitiated oxidation suppresses unproductive back-electron-transfer pathways, which are generally fast compared with substrate transformation, and because their decomposition products can provide additional reducing equivalents needed for some CO₂ reduction reactions, thus circumventing the one-photon/one-electron limit of molecular photosensitizers. Offsetting these practical advantages, however, is the fact that the stoichiometric consumption of conventional sacrificial donors in these reactions negates their energy-storing potential. In order for homogeneous systems to drive CO₂ reduction reactions that store energy, these sacrificial reagents must be replaced by a second catalytic cycle that extracts the reducing equivalents from a renewable source (16).Neumann and coworkers recently reported a photochemical system for the reduction of CO₂ to CO in which the reducing equivalents are derived from the oxidation of H₂ by colloidal platinum (15). This system, which contains a [Re^I(phen)(CO)₃L]⁺ (phen is 1,10-phenanthroline) CO₂ reduction catalyst linked with a polyoxometalate cluster, drives the reverse water–gas-shift reaction (RWGS) (CO₂ + H₂ → CO + H₂O), using light as the energy source, via the reactions shown in Eqs. 1–3. Unlike photochemical reactions that consume sacrificial donors, this energy-storing system [ΔH_f = 41.2 kJ⋅mol^–1 (17)] catalytically extracts renewable reducing equivalents that can be sourced to water.H₂→2e^? + 2H⁺[1]CO₂ + 2e^? + 2H⁺→CO + H₂O[2]CO₂ + H₂→CO + H₂O[3]The mechanistic, thermodynamic, and kinetic integration of two catalytic cycles with a chromophore is a general challenge that cuts across homogeneous molecular approaches to forming solar fuels from CO₂ and H₂O. A photochemical system for the RWGS reaction that used a homogeneous H₂ oxidation catalyst would both provide insights into the fundamental factors that govern this integration and opportunities to exert greater control over them than is possible with heterogeneous catalysts. Motivated by these possibilities, we report here the photochemistry of a homogeneous system composed of a photosensitizer, CO₂-reduction catalyst, and H₂-oxidation catalyst that, upon excitation with long-wavelength light (λ > 590 nm), yields a product state thermodynamically capable of accomplishing the RWGS reaction. The system operates via reductive quenching of the low-energy T₁ excited state of the common chromophore zinc tetraphenylporphyrin (ZnTPP) by a compound of the form Cp^RCr(CO)₃^– (1^–; Cp^R = η⁵-cyclopentadienyl), followed by thermal electron transfer from the product ZnTPP^– radical to a complex of the type Re(bpy-4,4′-R₂)(CO)₃Cl (2; bpy is 2,2′-bipyridyl). The electron-transfer-sensitized radical products of these reactions—Cp^RCr(CO)₃ (1•) and [Re(bpy-4,4′-R₂)(CO)₃Cl]^– (2^–)—can initiate the oxidation of H₂ and reduction of CO₂, respectively. The ligand substituents within each of these classes of compounds allow control over the driving forces and, thus, the rates of the productive (and unproductive) electron-transfer reactions available to the components. A comprehensive picture of the mechanism and kinetics of this system has been elucidated using time-resolved spectroscopic methods.     

Origin of enhanced water oxidation activity in an iridium single atom anchored on NiFe oxyhydroxide catalyst

The efficiency of the synthesis of renewable fuels and feedstocks from electrical sources is limited, at present, by the sluggish water oxidation reaction. Single-atom catalysts (SACs) with a controllable coordination environment and exceptional atom utilization efficiency open new paradigms toward designing high-performance water oxidation catalysts. Here, using operando X-ray absorption spectroscopy measurements with calculations of spectra and electrochemical activity, we demonstrate that the origin of water oxidation activity of IrNiFe SACs is the presence of highly oxidized Ir single atom (Ir^5.3+) in the NiFe oxyhydroxide under operating conditions. We show that the optimal water oxidation catalyst could be achieved by systematically increasing the oxidation state and modulating the coordination environment of the Ir active sites anchored atop the NiFe oxyhydroxide layers. Based on the proposed mechanism, we have successfully anchored Ir single-atom sites on NiFe oxyhydroxides (Ir_0.1/Ni₉Fe SAC) via a unique in situ cryogenic–photochemical reduction method that delivers an overpotential of 183 mV at 10 mA ⋅ cm⁻² and retains its performance following 100 h of operation in 1 M KOH electrolyte, outperforming the reported catalysts and the commercial IrO₂ catalysts. These findings open the avenue toward an atomic-level understanding of the oxygen evolution of catalytic centers under in operando conditions.

Efficient and cost-effective electrocatalysts play critical roles in energy conversion and storage and the societal pursuit of sustainable energy (1–3). The water oxidation reaction, also known as oxygen evolution reaction (OER), in particular, is an enabling process for diverse clean energy technologies, including water splitting (4–6), solar fuels (2), CO₂ reduction (7), and rechargeable metal–air batteries (8). Unfortunately, the kinetics of the OER are sluggish, which limits the power conversion efficiency and the overall efficiency.Very recently, higher valence transition metal ions such as Co⁴⁺ (9–11), Ni⁴⁺ (12–14), and Fe^4+/5+ (15, 16) generated through a potential-dependent deprotonation reaction have been incorporated into metal oxides/hydroxides, resulting in enhanced water oxidation activity. Incorporating precious metals such as Ir, Ru, and Pt is much less explored, but they offer greater opportunities due to their tendency to form single-atom sites. Computational work has predicted either direct substitution of Ni⁴⁺ and Fe⁴⁺ by Ir⁴⁺ and Ru⁴⁺ or that the metal would be close to its most stable +4 oxidation state based on the high stability of the rutile phase (17–20). On the other hand, the high activity of Sr-leached SrIrO₃/IrO_x and Li-removed Li_xIrO₃ catalysts (5, 21, 22) and a recent prediction of highly active and high-oxidation homogeneous water oxidation systems (23) indicate that increased oxidation may lead to improved activity if the active site can be stabilized under operating conditions.Single-atom catalysts (SACs) have offered an ideal system to precisely control the local coordination environments and oxidation states of the single-site centers (24–29). The single-atom nature of these active centers leads to well-defined coordination environments and enhanced metal–support interactions, which provide remarkable catalytic performance in a number of heterogeneous reactions, including in the water–gas shift reaction (24), heterogeneous reduction of CO₂ processes (27), CO oxidation (28), and oxygen reduction reaction (29). We adopted this strategy and sought to incorporate high-oxidation Ir metal sites into the support to enhance the water oxidation activity.Here, we developed an in situ cryogenic–photochemical reduction method for anchoring Ir single sites on the NiFe oxyhydroxide support. Density functional theory (DFT) calculations predict unusually stable IrO₆ octahedral SAC anchored atop the NiFe oxyhydroxide layers. In operando Ir L₃-edge X-ray absorption spectroscopy (XAS) combined with spectra simulations of Ir L₃-edge of such structures revealed high oxidation of Ir (+5.3) in the NiFe oxyhydroxides under operating conditions, in agreement with DFT predictions and spectra calculations. The highly oxidized Ir single-site exhibits exceptional OER performance, with a 183 mV overpotential at 10 mA ⋅ cm⁻², outperforming the precious metal oxide IrO₂. These findings are further corroborated by our calculations of the theoretical OER overpotentials, which show that increasing the Ir oxidation state leads to improved activity, and an overpotential of 0.184 V is obtained for the most oxidized NiFeIr:3O SAC. Such structural and compositional heterogeneity poses a key obstacle to unambiguously identifying the exact atomistic structure of the active sites and to further establishing a definitive correlation with the catalytic properties that can guide the subsequent design of future generations of SACs.     

INAUGURAL ARTICLE by a Recently Elected Academy Member:From the Cover: Nickel-borate oxygen-evolving catalyst that functions under benign conditions

Thin catalyst films with electrocatalytic water oxidation properties similar to those of a recently reported Co-based catalyst can be electrodeposited from dilute Ni²⁺ solutions in borate electrolyte at pH 9.2 (B_i). The Ni-B_i films can be prepared with precise thickness control and operate at modest overpotential providing an alternative to the Co catalyst for applications in solar energy conversion.     

Reaction mechanism of melatonin oxidation by reactive oxygen species in vitro

Melatonin (N-acetyl-5-hydroxytryptamine) is a pineal hormone widely known for its antioxidant properties, both in vivo and by direct capture of free radicals in vitro. Although some metabolites and oxidation products of melatonin have been identified, the molecular mechanism by which melatonin exerts its antioxidant properties has not been totally unravelled. This study investigated the reaction mechanism of oxidation of melatonin by radio-induced reactive oxygen species, generated by gamma radiolysis of water for aqueous solutions of melatonin (from 20 to 200 μm), in the presence or absence of molecular oxygen. The hydroxyl radical was found to be the unique species able to initiate the oxidation process, leading to three main products, e.g. N(1)-acetyl-N(2)-formyl-5-methoxykynurenin (AFMK), N(1)-acetyl-5-methoxykynurenin (AMK) and hydroxymelatonin (HO-MLT). The generation of AFMK and HO-MLT strongly depended on the presence of molecular oxygen in solution: AFMK was the major product in aerated solutions (84%), whereas HO-MLT was favoured in the absence of oxygen (86%). Concentrations of AMK remained quite low, and AMK was proposed to result from a chemical hydrolysis of AFMK in solution. A K-value of 1.1 × 10(-4) was calculated for this equilibrium. Both hydrogen peroxide and superoxide dismutase had no effect on the radio-induced oxidation of melatonin, in good accordance for the second case with the poor reactivity of the superoxide anion towards melatonin. Finally, a reaction mechanism was proposed for the oxidation of melatonin in vitro.     

The electrochemical approach to concerted proton—electron transfers in the oxidation of phenols in water

Establishing mechanisms and intrinsic reactivity in the oxidation of phenol with water as the proton acceptor is a fundamental task relevant to many reactions occurring in natural systems. Thanks to the easy measure of the reaction kinetics by the current and the setting of the driving force by the electrode potential, the electrochemical approach is particularly suited to this endeavor. Despite challenging difficulties related to self-inhibition blocking the electrode surface, experimental conditions were established that allowed a reliable analysis of the thermodynamics and mechanisms of the proton-coupled electron-transfer oxidation of phenol to be carried out by means of cyclic voltammetry. The thermodynamic characterization was conducted in buffer media whereas the mechanisms were revealed in unbuffered water. Unambiguous evidence of a concerted proton–electron transfer mechanism, with water as proton acceptor, was thus gathered by simulation of the experimental data with appropriately derived theoretical relationships, leading to the determination of a remarkably large intrinsic rate constant. The same strategy also allowed the quantitative analysis of the competition between the concerted proton–electron transfer pathway and an OH⁻-triggered stepwise pathway (proton transfer followed by electron transfer) at high pHs. Investigation of the passage between unbuffered and buffered media with the example of the PO₄H₂⁻/PO₄H²⁻ couple revealed the prevalence of a mechanism involving a proton transfer preceding an electron transfer over a PO₄H²⁻-triggered concerted process.     

High-resolution cryo-electron microscopy structure of photosystem II from the mesophilic cyanobacterium,Synechocystis sp. PCC 6803

Photosystem II (PSII) enables global-scale, light-driven water oxidation. Genetic manipulation of PSII from the mesophilic cyanobacterium Synechocystis sp. PCC 6803 has provided insights into the mechanism of water oxidation; however, the lack of a high-resolution structure of oxygen-evolving PSII from this organism has limited the interpretation of biophysical data to models based on structures of thermophilic cyanobacterial PSII. Here, we report the cryo-electron microscopy structure of PSII from Synechocystis sp. PCC 6803 at 1.93-Å resolution. A number of differences are observed relative to thermophilic PSII structures, including the following: the extrinsic subunit PsbQ is maintained, the C terminus of the D1 subunit is flexible, some waters near the active site are partially occupied, and differences in the PsbV subunit block the Large (O1) water channel. These features strongly influence the structural picture of PSII, especially as it pertains to the mechanism of water oxidation.

Photosystem II (PSII) is a multisubunit membrane protein complex found in oxygenic phototrophs. It is the only global-scale catalyst for solar fuel production (1). Thus, understanding its enzymatic function is relevant to a variety of fields including synthetic photocatalysis, crop optimization, biofuel production, and evolutionary biology. PSII catalyzes redox processes that result in the transfer of electrons from water on the “donor side” (lumenal side) to lower potential plastoquinone on the “acceptor side” (stromal side). The active site of PSII contains an inorganic metallocofactor, a Mn₄CaO₅ cluster termed the oxygen-evolving complex (OEC) that is coordinated primarily by one of the core subunits, D1. The mechanism of water oxidation at the OEC proceeds through a series of five intermediate storage states, or S-states, S₀ through S₄, referred to as the Kok cycle, which have been the subject of extensive study (2–4). Various water channels extend away from the OEC toward the lumen and are involved in substrate water delivery, proton transfer, and oxygen release (2–5). In cyanobacteria, the extrinsic subunits bound to the lumenal side, PsbO, PsbQ, PsbU, and PsbV, play different roles in stabilizing the OEC and protecting it from reductants that can disrupt water oxidation, and they also contribute to water channel formation (6, 7). In plants and algae, analogous but evolutionarily distinct extrinsic subunits are bound. Despite many years of interdisciplinary efforts aimed at understanding the details of water oxidation, many fundamental aspects remain unclear due to the highly complicated nature of the reaction mechanism.The ability to perform site-directed mutagenesis has contributed greatly to our understanding of PSII, especially the mechanism of water oxidation. A genetic system was first reported in the late 1980s in a mesophilic cyanobacterium, Synechocystis sp. PCC 6803 (hereafter Synechocystis 6803) (8), which is a convenient model organism to study due to its ability to grow in the absence of PSII when supplemented with glucose. Given the ease of genetic transformation (9, 10), many site-directed mutated PSII complexes have been investigated (11–13). PSII point mutants have been used in biophysical experiments such as Fourier transform infrared spectroscopy (FTIR) (14, 15), electron paramagnetic resonance spectroscopy (16), mass spectrometry (17), and more to assess the role of individual amino acids in PSII function. Equally as informative have been studies of PSII using structural approaches such as X-ray crystallography (4) and more recently cryo-electron microscopy (cryo-EM) (18), although no structures have yet been solved of PSII with point mutations.All active cyanobacterial PSII structures reported to date have been solved from the thermophiles Thermosynechococcus vulcanus and Thermosynechococcus elongatus, which are nearly identical in sequence and structure. Typically, it is assumed that the structure and function of PSII are highly conserved between mesophilic and thermophilic cyanobacteria. As a result, biophysical data obtained from PSII isolated from mesophilic cyanobacteria are commonly interpreted using molecular structures of PSII obtained from thermophilic cyanobacteria. This approach may be problematic because membrane proteins from mesophilic and thermophilic organisms are generally known to exhibit differences in molecular interactions (19), and there are obvious sequence differences between PSII subunits from mesophilic and thermophilic cyanobacteria that imply structural and functional variation (SI Appendix, Fig. S1 and Table S1). Thus, a glaring roadblock in the study of PSII is the lack of a molecular structure from the easily transformable mesophilic cyanobacterium, Synechocystis 6803, that is the source of so much biochemical and biophysical data.Here, we present the cryo-EM structure of PSII from Synechocystis 6803 at 1.93-Å resolution. This structure is compared with previously solved structures of PSII, especially a recent cryo-EM structure from T. vulcanus (20). The extrinsic subunit PsbQ that is missing in other cyanobacterial PSII structures is bound at a site nearly identical to that of PsbQ in higher plants and algae. The OEC exhibits differences in its coordination by the C terminus of the D1 subunit and variable positions of some nearby amino acids and waters are observed. The Large water channel (also known as the O1 channel) present in PSII from thermophilic cyanobacteria (21) is blocked in the Synechocystis 6803 PSII structure, owing to differences in the PsbV subunit. To assess radiation damage to the sample, a low-dose Synechocystis 6803 PSII cryo-EM structure was determined at a resolution of 2.01 Å, which shows negligible differences compared with the full-dose structure. The high-resolution structure presented here provides a view of fully assembled PSII from a mesophilic cyanobacterium. This provides a model from which past and future biochemical and biophysical data can be more accurately interpreted and it establishes a method to solve molecular structures of PSII with single amino acid substitutions that will address questions regarding PSII activity.     

Pathway for Mn-cluster oxidation by tyrosine-Z in the S2 state of photosystem II

Water oxidation in photosynthetic organisms occurs through the five intermediate steps S₀–S₄ of the Kok cycle in the oxygen evolving complex of photosystem II (PSII). Along the catalytic cycle, four electrons are subsequently removed from the Mn₄CaO₅ core by the nearby tyrosine Tyr-Z, which is in turn oxidized by the chlorophyll special pair P680, the photo-induced primary donor in PSII. Recently, two Mn₄CaO₅ conformations, consistent with the S₂ state (namely, S2A and S2B models) were suggested to exist, perhaps playing a different role within the S₂-to-S₃ transition. Here we report multiscale ab initio density functional theory plus U simulations revealing that upon such oxidation the relative thermodynamic stability of the two previously proposed geometries is reversed, the S2B state becoming the leading conformation. In this latter state a proton coupled electron transfer is spontaneously observed at ∼100 fs at room temperature dynamics. Upon oxidation, the Mn cluster, which is tightly electronically coupled along dynamics to the Tyr-Z tyrosyl group, releases a proton from the nearby W1 water molecule to the close Asp-61 on the femtosecond timescale, thus undergoing a conformational transition increasing the available space for the subsequent coordination of an additional water molecule. The results can help to rationalize previous spectroscopic experiments and confirm, for the first time to our knowledge, that the water-splitting reaction has to proceed through the S2B conformation, providing the basis for a structural model of the S₃ state.For 2.5 Gy photosynthetic organisms have used the photosystem II complex (PSII) to capture light energy from the sun and convert it into chemical energy stored within energy-rich carbohydrates (1). The water oxidation reaction, occurring in the reaction center of PSII, represents the central step of the natural photosynthetic process, leading to the formation of molecular oxygen and hydrogen equivalents. A deep understanding of the photosynthetic water-splitting mechanism may serve as a valuable source of inspiration for the development of artificial devices able to store solar energy in environmentally friendly fuels, such as molecular hydrogen (2–6). The active site of the PSII enzyme, where the water-splitting reaction takes place, consists of a core of four Mn ions and one Ca ion connected together through μ-oxo bridges in a cubane-like aggregate (7). Water oxidation proceeds through five sequential S₀–S₄ steps known as the Kok cycle (8). At each step of the catalytic cycle the absorption of photons turns out the oxidation of the tyrosyl group of a nearby tyrosine (i.e., Tyr161, also known as Tyr-Z in the D1 subunit of PSII), which acts as an intermediate in the electron transfer between the Mn₄CaO₅ cluster and the primary donor P680 (9, 10).The molecular structure for the different states of the Kok cycle was largely investigated in the past decades by extended X-ray absorption fine structure experiments (11–14) and X-ray crystallography (15–17), thus revealing atomic details at an increasing level of accuracy (7). In parallel the electronic and magnetic properties, characterizing steps of the catalytic cycle, were investigated by EPR experiments (18–24), with particular attention to the well-characterized S₂ state (25). The combination of this large number of experimental results within a computational modeling framework gave the opportunity to understand some of the atomic details underlying the water-splitting reaction in the oxygen-evolving complex (OEC) (26–35). Recently, Pantazis et al. (36) have suggested the presence of two possible interconvertible structures representative of the S₂ state. The two models, namely, the S2A state, characterized by a S = 1/2 spin ground state, and the S2B state, characterized by a S = 5/2 spin ground state (Fig. 1), can explain the presence of the two distinct EPR signals revealed at cryogenic temperatures (i.e., a multiline signal indicative of a ground state characterized by a spin S = 1/2 and a second signal at g = 4.1 consistent with a spin S = 5/2). In a recent work (37) we characterized by extensive quantum mechanics/molecular mechanics (QM/MM) ab initio simulations the free-energy profiles for the interconversion between the two above-mentioned conformations, suggesting that the transition from the S₂ to S₃ state should proceed passing first by the S2A and subsequently through the S2B state. Still, clear evidence confirming such a hypothesis is missing.Open in a separate windowFig. 1.Ab initio QM/MM model of photosystem II. (Right) The QM region, consisting of 224 atoms, is shown in balls and sticks representation. (Upper Left) A selection of the the most important residues and distances involved in the oxidation of the Mn₄CaO₅ cluster by the radical Tyr-Z are sketched. (Lower Left) Representation of the two investigated conformations S2A and S2B.Here, using the same approach previously adopted, we characterized different (spin) energy surfaces along the interconversion path between the S2A and S2B states after the removal of one electron from the QM region (Fig. 1). QM/MM molecular dynamic simulations were additionally carried out for the two models in their respective spin ground state: the low spin (LS) state for the S2A model and the high spin (HS) state for the S2B model. The present results show for the first time, to our knowledge, the occurrence of a proton coupled electron transfer (PCET) in the S2B state resulting in the oxidation of the Mn4 ion by the tyrosyl group of Tyr-Z.     

Alternating electron and proton transfer steps in photosynthetic water oxidation

Water oxidation by cyanobacteria, algae, and plants is pivotal in oxygenic photosynthesis, the process that powers life on Earth, and is the paradigm for engineering solar fuel–production systems. Each complete reaction cycle of photosynthetic water oxidation requires the removal of four electrons and four protons from the catalytic site, a manganese–calcium complex and its protein environment in photosystem II. In time-resolved photothermal beam deflection experiments, we monitored apparent volume changes of the photosystem II protein associated with charge creation by light-induced electron transfer (contraction) and charge-compensating proton relocation (expansion). Two previously invisible proton removal steps were detected, thereby filling two gaps in the basic reaction-cycle model of photosynthetic water oxidation. In the S₂ → S₃ transition of the classical S-state cycle, an intermediate is formed by deprotonation clearly before electron transfer to the oxidant (). The rate-determining elementary step (τ, approximately 30 µs at 20 °C) in the long-distance proton relocation toward the protein–water interface is characterized by a high activation energy (E_a = 0.46 ± 0.05 eV) and strong H/D kinetic isotope effect (approximately 6). The characteristics of a proton transfer step during the S₀ → S₁ transition are similar (τ, approximately 100 µs; E_a = 0.34 ± 0.08 eV; kinetic isotope effect, approximately 3); however, the proton removal from the Mn complex proceeds after electron transfer to . By discovery of the transient formation of two further intermediate states in the reaction cycle of photosynthetic water oxidation, a temporal sequence of strictly alternating removal of electrons and protons from the catalytic site is established.     

人工水和自然水环境中嗜肺军团菌mip基因分型

目的 比较人工水体与自然水体环境中军团菌的mip基因差异.方法 将2003-2008年间广州和新会两市水样中分离的121株嗜肺军团菌分为人工与自然水体环境分离株两组,分别对其mip基因进行系统进化树重建,基因多态性分析,分子方差分析与中性检验,并比较两种分离来源菌株的mip基因差异.结果 121株嗜肺军团菌mip基因按分离环境来源不均匀地分布在系统进化树的拓扑分支.人工水体环境分离株共有8个等位基因型,以等位基因1型为优势群,比例55.36％,自然水体环境分离株有11个等位基因型,以等位基因28型为优势群,比例40％.人工水体环境分离株的核苷酸多态性与平均核苷酸差异大于自然水体环境分离株.分子方差分析结果显示两种环境分离株mip基因差异占总体差异的14.43％.中性检验证明人工水体环境分离株的mip3’末端处,Tajima检验P＜0.05,D值显著大于0.而自然环境分离株在此区域未见显著性差异.结论 嗜肺军团菌mip基因在人工与自然水体环境分离株之间存在显著差异,这种差异可能来源于人工水体环境对嗜肺军团菌种群大小的影响或平衡选择作用.     

Visible photoelectrochemical water splitting into H2 and O2 in a dye-sensitized photoelectrosynthesis cell

A hybrid strategy for solar water splitting is exploited here based on a dye-sensitized photoelectrosynthesis cell (DSPEC) with a mesoporous SnO₂/TiO₂ core/shell nanostructured electrode derivatized with a surface-bound Ru(II) polypyridyl-based chromophore–catalyst assembly. The assembly, [(4,4’-(PO₃H₂)₂bpy)₂Ru(4-Mebpy-4’-bimpy)Ru(tpy)(OH₂)]⁴⁺ ([Ru_a^II-Ru_b^II-OH₂]⁴⁺, combines both a light absorber and a water oxidation catalyst in a single molecule. It was attached to the TiO₂ shell by phosphonate-surface oxide binding. The oxide-bound assembly was further stabilized on the surface by atomic layer deposition (ALD) of either Al₂O₃ or TiO₂ overlayers. Illumination of the resulting fluorine-doped tin oxide (FTO)|SnO₂/TiO₂|-[Ru_a^II-Ru_b^II-OH₂]⁴⁺(Al₂O₃ or TiO₂) photoanodes in photoelectrochemical cells with a Pt cathode and a small applied bias resulted in visible-light water splitting as shown by direct measurements of both evolved H₂ and O₂. The performance of the resulting DSPECs varies with shell thickness and the nature and extent of the oxide overlayer. Use of the SnO₂/TiO₂ core/shell compared with nanoITO/TiO₂ with the same assembly results in photocurrent enhancements of ∼5. Systematic variations in shell thickness and ALD overlayer lead to photocurrent densities as high as 1.97 mA/cm² with 445-nm, ∼90-mW/cm² illumination in a phosphate buffer at pH 7.Although promising, significant challenges remain in the search for successful strategies for artificial photosynthesis by water splitting into oxygen and hydrogen or reduction of CO₂ to reduced forms of carbon (1–5). In a dye-sensitized photoelectrosynthesis cell (DSPEC), a wide band gap, nanoparticle oxide film, typically TiO₂, is derivatized with a surface-bound molecular assembly or assemblies for light absorption and catalysis (6–8). In a DSPEC, visible light is absorbed by a chromophore, initiating a series of events that culminate in water splitting: injection, intraassembly electron transfer, catalyst activation, and electron transfer to a cathode or photocathode for H₂ production. Sun and coworkers have recently demonstrated visible-light–driven water splitting with a coloading approach combining Ru(II) polypyridyl-based light absorbers and catalysts on TiO₂ (9). The efficiency of DSPEC devices is dependent on interfacial dynamics and competing kinetic processes. A major limiting factor is the requirement for accumulating multiple oxidative equivalents at a catalyst site to meet the 4e⁻/4H⁺ demands for oxidizing water to dioxygen (2H₂O - 4e⁻ - 4H⁺ → O₂) in competition with back electron transfer of injected electrons to the oxidized assembly.One approach to achieving structural control of local electron transfer dynamics at the oxide interface in dye-sensitized devices is by use of nanostructured core/shell electrodes (10–12). In this approach, a mesoporous network of nanoparticles is uniformly coated with a thin oxide overlayer prepared by atomic layer deposition (ALD). We have used core/shell electrodes to demonstrate benzyl alcohol dehydrogenation (13). This approach has also been used to enhance the efficiency of dye-sensitized solar cells (14, 15). Recently, we described the use of a core/shell consisting of an inner core of a nanoparticle transparent conducting oxide, tin-doped indium oxide (nanoITO), and a thin outer shell of TiO₂ for water splitting by visible light (16). Derivatization of the nanoITO/TiO₂ core/shell electrode by surface binding of the chromophore–catalyst assembly, [(4,4’-(PO₃H₂)₂bpy)₂Ru_a(4-Mebpy-4’-bimpy)Ru_b(tpy)(OH₂)]⁴⁺ (1; -[Ru_a^II-Ru_b^II-OH₂]⁴⁺) shown in Fig. 1A, provided the basis for a photoanode in a DSPEC application with a Pt cathode for H₂ generation with a small applied bias in an acetate buffer at pH 4.6.Open in a separate windowFig. 1.(A) Chemical structure of chromophore–catalyst assembly 1, -[Ru_a^II-Ru_b^II-OH₂]⁴⁺. (B) TEM depicting a core/shell nanostructure from 75 ALD cycles of TiO₂ deposited onto SnO₂ films on FTO glass (FTO|SnO₂/TiO₂(4.5 nm)|). (C) Cartoon depicting an ALD core/shell electrode surface with and without ALD overlayer stabilization of a surface-bound assembly.Application of the core/shell structure led to a greatly enhanced efficiency for water splitting compared with mesoscopic, nanoparticle TiO₂ but the per-photon absorbed efficiency of the resulting DSPEC was relatively low and problems arose from long-term instability due to loss of the assembly from the oxide surface in the acetate buffer at pH 4.6. The latter is problematic because the rate of water oxidation is enhanced by added buffer bases, conditions that also enhance the rate of water oxidation (5, 17–24).Here, we report a second-generation DSPEC based on a core/shell photoanode. It features both greatly enhanced efficiencies for visible-light–driven water splitting and stabilization of surface binding by the assembly. Enhanced efficiencies come from the use of a SnO₂ core in a SnO₂/TiO₂ core/shell structure. SnO₂ has a conduction band potential (E_CB) more positive than TiO₂ by ∼0.4 V. Once injection and electron transfer to the SnO₂ core has occurred, an internal potential gradient at the SnO₂/TiO₂ interface is established, inhibiting back electron transfer.In the second-generation DSPEC, ALD is also used to stabilize oxide surface binding by the phosphonate-derivatized assembly. ALD deposition of overlayers of TiO₂ or Al₂O₃ has been shown to greatly enhance surface stability toward hydrolysis even in strongly basic solutions (25, 26). We show here, for assembly 1 surface-bound to SnO₂/TiO₂, that ALD overlayers of TiO₂ or Al₂O₃ provide both long-term stabilization on the oxide surface at pH 7 in a phosphate buffer, and, as a bonus, incrementally enhanced efficiencies for water splitting (23).The underlying strategy behind the use of ALD for both core/shell structure and stabilized surface binding is illustrated in Fig. 1C. Detailed information about the mechanism and rate of water oxidation by the surface-bound assembly is available from a previous publication (27).     

Similarities of artificial photosystems by ruthenium oxo complexes and native water splitting systems

The nature of chemical bonds of ruthenium(Ru)–quinine(Q) complexes, mononuclear [Ru(trpy)(3,5-t-Bu₂Q)(OH₂)](ClO₄)₂ (trpy = 2,2^′:6^′,2^′′-terpyridine, 3,5-di-tert-butyl-1,2-benzoquinone) (1), and binuclear [Ru₂(btpyan)(3,6-di-Bu₂Q)₂(OH₂)]²⁺ (btpyan = 1,8-bis(2,2^′:6^′,2^′′-terpyrid-4^′-yl)anthracene, 3,6-t-Bu₂Q = 3,6-di-tert-butyl-1,2-benzoquinone) (2), has been investigated by broken-symmetry (BS) hybrid density functional (DFT) methods. BS DFT computations for the Ru complexes have elucidated that the closed-shell structure (2b) Ru(II)–Q complex is less stable than the open-shell structure (2bb) consisting of Ru(III) and semiquinone (SQ) radical fragments. These computations have also elucidated eight different electronic and spin structures of tetraradical intermediates that may be generated in the course of water splitting reaction. The Heisenberg spin Hamiltonian model for these species has been derived to elucidate six different effective exchange interactions (J) for four spin systems. Six J values have been determined using total energies of the eight (or seven) BS solutions for different spin configurations. The natural orbital analyses of these BS DFT solutions have also been performed in order to obtain natural orbitals and their occupation numbers, which are useful for the lucid understanding of the nature of chemical bonds of the Ru complexes. Implications of the computational results are discussed in relation to the proposed reaction mechanisms of water splitting reaction in artificial photosynthesis systems and the similarity between artificial and native water splitting systems.     

Experimental demonstration of radicaloid character in a RuV=O intermediate in catalytic water oxidation

Water oxidation is the key half reaction in artificial photosynthesis. An absence of detailed mechanistic insight impedes design of new catalysts that are more reactive and more robust. A proposed paradigm leading to enhanced reactivity is the existence of oxyl radical intermediates capable of rapid water activation, but there is a dearth of experimental validation. Here, we show the radicaloid nature of an intermediate reactive toward formation of the O-O bond by assessing the spin density on the oxyl group by Electron Paramagnetic Resonance (EPR). In the study, an ¹⁷O-labeled form of a highly oxidized, short-lived intermediate in the catalytic cycle of the water oxidation catalyst cis,cis-[(2,2-bipyridine)₂(H₂O)Ru^IIIORu^III(OH₂)(bpy)₂]⁴⁺ was investigated. It contains Ru centers in oxidation states [4,5], has at least one Ru^V = O unit, and shows |A_xx| = 60G ¹⁷O hyperfine splittings (hfs) consistent with the high spin density of a radicaloid. Destabilization of π-bonding in the d³ Ru^V = O fragment is responsible for the high spin density on the oxygen and its high reactivity.     

Effect of lipid oxidation on the regulation of glucose utilization in obese patients

The effect of changes in lipid oxidation on glucose utilization (storage and oxidation) was studied in seven nondiabetic obese patients. They participated in three protocols in which: (1) Intralipid (to raise plasma FFA concentrations), (2) -pyridylcarbinol [a precursor of nicotinic acid, to lower plasma free fatty acids (FFA) concentrations], or (3) isotonic saline were infused over 2 h. Thereafter, these infusions were discontinued, and a 2-h cuglycemic, hyperinsulinemic clamp was performed to measure glucose uptake. All studies were carried out in combination with indirect calorimetry to measure oxidative and nonoxidative glucose disposal (glucose storage). The high plasma FFA concentrations (1024±57 mol/l) and lipid oxidation rates (1.1±0.1 mg/kg·min) found at the end of the Intralipid infusion and the low plasma FFA concentrations (264±26 mol/l and lipid oxidation rates 0.7±0.1 mg/kg·min) found at the end of the -pyridylcarbinol infusions resulted in significantly different rates of total and nonoxidative glucose disposal during the insulin clamp. The values were 2.6±0.6 mg/kg·min after Intralipid and 4.1±1.0 mg/kg·min after -pyridylcarbinol for total glucose disposal, and 0.4±0.4 and 1.6±0.8, respectively for nonoxidative glucose disposal. In conclusion, these observations show that changes in lipid oxidation rates preceding a glucose load influence glucose disposal and glycogen storage in obese subjects.