Ammonia Binding Research Articles

Stability and Structure of Potentially Atmospherically Relevant Glycine Ammonium Bisulfate Clusters.

New particle formation (NPF) is the process by which trace atmospheric acids and bases cluster and grow into particles that ultimately impact climate. Sulfuric acid concentration drives NPF, but nitrogen-containing bases promote the formation of more stable clusters via salt bridge formation. Recent computational efforts have suggested that amino acids can enhance NPF, predicting that they can stabilize new particles via multiple protonation sites, but there has yet to be experimental validation of these predictions. We used mass spectrometry and infrared spectroscopy to study the structure and stability of cationic clusters composed of glycine, sulfuric acid, and ammonia. When collisionally activated, clusters were significantly more likely to eliminate ammonia or sulfuric acid than glycine, while quantum chemical calculations predicted lower binding free energies for ammonia but similar binding free energies for glycine and sulfuric acid. These calculations predicted several low-energy structures, so we compared experimental and computed vibrational spectra to attempt to validate the computationally predicted minimum energy structure. Unambiguous identification of the experimental structure by comparison to these calculations was made difficult by the complexity of the experimental spectra and the fact that the identity of the computed lowest-energy structure depended strongly on temperature. If their vapors are present, amino acids are likely to be enriched in new particles by displacing more weakly bound ammonia, similar to the behavior of other atmospheric amines. The carboxylic acid groups were found to preferentially interact with other carboxylic acids, suggesting incipient organic/inorganic phase separation even at these small sizes.

Comprehensive Evaluation of Models for Ammonia Binding to the Oxygen Evolving Complex of Photosystem II.

The identity and insertion pathway of the substrate oxygen atoms that are coupled to dioxygen by the oxygen-evolving complex (OEC) remains a central question toward understanding Nature's water oxidation mechanism. In several studies, ammonia has been used as a small "water analogue" to elucidate the pathway of substrate access to the OEC and to aid in determining which of the oxygen ligands of the tetramanganese cluster are substrates for O-O bond formation. On the basis of structural and spectroscopic investigations, five first-sphere binding modes of ammonia have been suggested, involving either substitution of an existing H2O/OH-/O2- group or addition as an extra ligand to a metal ion of the Mn4CaO5 cluster. Some of these modes, specifically the ones involving substitution, have already been subject to spectroscopy-oriented quantum chemical investigations, whereas more recent suggestions that postulate the addition of ammonia have not been examined so far with quantum chemistry for their agreement with spectroscopic data. Herein, we use a common structural framework and theoretical methodology to evaluate structural models of the OEC that represent all proposed modes of first-sphere ammonia interaction with the OEC in its S2 state. Criteria include energetic, magnetic, kinetic, and spectroscopic properties compared against available experimental EPR, ENDOR, ESEEM, and EDNMR data. Our results show that models featuring ammonia replacing one of the two terminal water ligands on Mn4 align best with experimental data, while they definitively exclude substitution of a bridging μ-oxo ligand as well as incorporation of ammonia as a sixth ligand on Mn1 or Mn4.

Validation Methods for the Ammonia Removal of an Air Scrubber on a Poultry House Using the Acid Use and the Process Water Nitrogen Balance

Highlights Acid use in an acid air scrubber predicts the ammonia removal of an air scrubber. Nitrogen balance of process and discharge water in an air scrubber predicts ammonia removal of an air scrubber. Long-term monitoring of acid use and/or the nitrogen balance can validate the ammonia reduction of an air scrubber. The two validation methods can support local authorities in granting environmental permits and enforcement. Abstract. Ensured ammonia reduction by novel technologies and guaranteed low ammonia emission from livestock houses is essential for the granting and monitoring of legal permits. This study focuses on methods to underpin the long-term effectiveness of an air scrubber and was done in a commercial poultry house. The measured ammonia removal by an air scrubber with ammonia sensors (method 1) was compared to the acid use for ammonia binding (method 2) and the nitrogen balance over the process water (method 3). Also, the effect of the length of the time period used for the comparison was evaluated, as was the effect of the spatial variation of the ammonia concentration in the house. The overall weekly data showed a clear linear relation between method 1 on the one hand and methods 2 and 3 on the other hand, with regression coefficients close to 1 and a variation of +/- 20% on a 4-weekly basis. The 4-weekly data showed a clearly reduced variation compared to weekly data, due to leveling out of the impact of irregular time events like acid dosage and water discharge. The barn emission assessed by method 1 was significantly (12%) influenced by the spatial aerial concentration differences of ammonia in the house. Taking this into account, method 2 overestimated the emission reduction of method 1 with 16.8% and method 3 was in good agreement with method 1. For method 2, a revised value for acid use per kg of ammonia binding needs to be substantiated in further research. Possible measurement errors influenced calculated ammonia removal with a maximum of +/-2%, but the relative effect on remaining emissions was larger with +/- 15% for method 3. This study shows that monitoring acid use and/or the nitrogen balance can validate the ammonia reduction of air scrubbers and the remaining ammonia emission and support local authorities in granting environmental permits and their enforcement. Keywords: Acid, Air scrubber, Ammonia, Nitrogen balance, Poultry.

Extreme accumulation of ammonia on electroreduced mackinawite: An abiotic ammonia storage mechanism in early ocean hydrothermal systems.

An increasing amount of evidence suggests that early ocean hydrothermal systems were sustained sources of ammonia, an essential nitrogen species for prebiotic synthesis of life's building blocks. However, it remains a riddle how the abiotically generated ammonia was retained at the vent-ocean interface for the subsequent chemical evolution. Here, we demonstrate that, under simulated geoelectrochemical conditions in early ocean hydrothermal systems ([Formula: see text][Formula: see text] V versus the standard hydrogen electrode), mackinawite gradually reduces to zero-valent iron ([Formula: see text]), generating interlayer [Formula: see text] sites. This reductive conversion leads to an up to 55-fold increase in the solid/liquid partition coefficient for ammonia, enabling over 90% adsorption of 1 mM ammonia in 1 M NaCl at neutral pH. A coordinative binding of ammonia on the interlayer [Formula: see text] sites was computed to be the major mechanism of selective ammonia adsorption. Mackinawite is a ubiquitous sulfide precipitate in submarine hydrothermal systems. Given its reported catalytic function in amination, the extreme accumulation of ammonia on electroreduced mackinawite should have been a crucial initial step for prebiotic nitrogen assimilation, paving the way to the origin of life.

Theoretical Distribution of the Ammonia Binding Energy at Interstellar Icy Grains: A New Computational Framework.

The binding energies (BE) of molecules on the interstellar grains are crucial in the chemical evolution of the interstellar medium (ISM). Both temperature-programmed desorption (TPD) laboratory experiments and quantum chemistry computations have often provided, so far, only single values of the BE for each molecule. This is a severe limitation, as the ices enveloping the grain mantles are structurally amorphous, giving rise to a manifold of possible adsorption sites, each with different BEs. However, the amorphous ice nature prevents the knowledge of structural details, hindering the development of a common accepted atomistic icy model. In this work, we propose a computational framework that closely mimics the formation of the interstellar grain mantle through a water by water accretion. On that grain, an unbiased random (but well reproducible) positioning of the studied molecule is then carried out. Here we present the test case of NH3, a ubiquitous species in the molecular ISM. We provide the BE distribution computed by a hierarchy approach, using the semiempirical xTB-GFN2 as a low-level method to describe the whole icy cluster in combination with the B97D3 DFT functional as a high-level method on the local zone of the NH3 interaction. The final ZPE-corrected BE is computed at the ONIOM(DLPNO-CCSD(T)//B97D3:xTB-GFN2) level, ensuring the best cost/accuracy ratio. The main peak of the predicted NH3 BE distribution is in agreement with experimental TPD and computed data in the literature. A second broad peak at very low BE values is also present, which has never been detected before. It may provide the solution to a longstanding puzzle about the presence of gaseous NH3 also observed in cold ISM objects.

Ammonia stimulates SCAP/Insig dissociation and SREBP-1 activation to promote lipogenesis and tumour growth.

Tumorigenesis is associated with elevated glucose and glutamine consumption, but how cancer cells can sense their levels to activate lipid synthesis is unknown. Here, we reveal that ammonia, released from glutamine, promotes lipogenesis via activation of sterol regulatory element-binding proteins (SREBPs), endoplasmic reticulum-bound transcription factors that play a central role in lipid metabolism. Ammonia activates the dissociation of glucose-regulated, N-glycosylated SREBP-cleavage-activating protein (SCAP) from insulin-inducible gene protein (Insig), an endoplasmic reticulum-retention protein, leading to SREBP translocation and lipogenic gene expression. Notably, 25-hydroxycholesterol blocks ammonia to access its binding site on SCAP. Mutating aspartate D428 to alanine prevents ammonia binding to SCAP, abolishes SREBP-1 activation and suppresses tumour growth. Our study characterizes the unknown role, opposite to sterols, of ammonia as a key activator that stimulates SCAP-Insig dissociation and SREBP-1 activation to promote tumour growth and demonstrates that SCAP is a critical sensor of glutamine, glucose and sterol levels to precisely control lipid synthesis.

Influence of Gypsum-Containing Waste on Ammonia Binding in Animal Waste Composting

The possibility of using gypsum-containing waste–citrogypsum, which is a by-product of the chemical biosynthesis of citric acid, was considered as an additive for composting poultry manure from poultry farms that practice litter-free poultry keeping. The research was carried out on an experimental batch of 1500 tons. The production of the batch was carried out by mixing citrogypsum with a moisture content of 30% and litterless chicken manure with a moisture content of no more than 80% in a ratio of 1:2. The resulting mixture was placed on an open landfill in piles 3 m wide, 1 m high and 400 m long and was mixed twice with a compost turner. Further processing consisted of mixing the mass once every seven days. The controlled parameters were changes in humidity, temperature, pH and nitrogen content during composting. In the course of the experiment, it was found that the introduction of citrogypsum into the composition of the compost helps to optimize the moisture, temperature and pH of the mixture, and a decrease in ammonia emission to 87% was recorded, with an increase in nitrogen content of 2.4 times compared to the initial value. It was concluded that citrogypsum can be used in composting poultry waste to reduce volatilization of ammonia and preserve nutrients.

Low-Coordinate Iron Chalcogenolates and Their Complexes with Diethyl Ether and Ammonia.

Treatment of Fe{N(SiMe3)2}2 with 2 equiv of the appropriate phenol or thiol affords the dimers {Fe(OC6H2-2,6-But2-4-Me)2}2 (1) and {Fe(OC6H3-2,6-But2)2}2 (2) or the monomeric Fe{SC6H3-2,6-(C6H3-2,6-Pri2)2}2 (3) in moderate to excellent yields. Recrystallization of 1 and 2 from diethyl ether gives the corresponding three-coordinate ether complexes Fe(OC6H3-2,6-But2-4-Me)2(OEt2) (4) and Fe(OC6H3-2,6-But2)2(OEt2) (5). In contrast, no diethyl ether complex is formed by the dithiolate 3. The 1H NMR spectra of 4 and 5 show equilibria between the ether complexes and the base-free dimers. A comparison of these spectra with those of the dimeric 1 and 2 allows an unambiguous assignment of the paramagnetically shifted signals. Treatment of 1 with excess ammonia gives the tetrahedral diammine Fe(OC6H2-2,6-But2-4-Me)2(NH3)2 (6). Ammonia is strongly coordinated in 6, with no apparent loss of ammine ligand either in solution or upon heating under low pressure. In contrast, significantly weaker ammonia coordination is observed when dithiolate 3 is treated with excess ammonia, which gives the diammine Fe{SC6H3-2,6-(2,6-Pri2-C6H3)2}2(NH3)2 (7). Complex 7 readily loses ammonia either in solution or under reduced pressure to give the monoammine complex Fe{SC6H3-2,6-(2,6-Pri2-C6H3)2}2(NH3) (8). The weak binding of ammonia by iron thiolate 7 reflects the likely behavior of the proposed iron-sulfur active site in nitrogenases, where release of ammonia is required to close the catalytic cycle.

Enzyme-Substrate-Cofactor Dynamical Networks Revealed by High-Resolution Field Cycling Relaxometry.

The remarkable power and specificity of enzyme catalysis rely on the dynamic alignment of the enzyme, substrates, and cofactors, yet the role of dynamics has usually been approached from the perspective of the protein. We have been using an underappreciated NMR technique, subtesla high-resolution field cycling 31P NMR relaxometry, to investigate the dynamics of the enzyme-bound substrates and cofactor on guanosine-5'-monophosphate reductase (GMPR). GMPR forms two dead end, yet catalytically competent, complexes that mimic distinct steps in the catalytic cycle: E·IMP·NADP+ undergoes a partial hydride transfer reaction, while E·GMP·NADP+ undergoes a partial deamination reaction. A different cofactor conformation is required for each partial reaction. Here we report the effects of mutations designed to perturb cofactor conformation and ammonia binding with the goal of identifying the structural features that contribute to the distinct dynamic signatures of the hydride transfer and deamination complexes. These experiments suggest that Asp129 is a central cog in a dynamic network required for both hydride transfer and deamination. In contrast, Lys77 modulates the conformation and mobility of substrates and cofactors in a reaction-specific manner. Thr105 and Tyr318 are part of a deamination-specific dynamic network that includes the 2'-OH of GMP. These residues have comparatively little effect on the dynamic properties of the hydride transfer complex. These results further illustrate the potential of high-resolution field cycling NMR relaxometry for the investigation of ligand dynamics. In addition, exchange experiments indicate that NH3/NH4+ has a high affinity for the deamination complex but a low affinity for the hydride transfer complex, suggesting that the movement of ammonia may gate the cofactor conformational change. Collectively, these experiments reinforce the view that the enzyme, substrates, and cofactor are linked in intricate, reaction-specific, dynamic networks and demonstrate that distal portions of the substrates and cofactors are critical features in these networks.

Influence of different types of bedding material on the prevalence of pododermatitis in rabbits

Pododermatitis is a disorder with a complex etiology, because potentially determined by various factors such as posture, nutrition and genetics. In the case of presence of pododermal lesions, it is often noticed that rabbits go to eat at the bowl less frequently, but an evident weight loss is not always perceptible. In fact, similarly to poultry, overweight animals seem to be prone to the onset of pododermatitis. An experiment was carried out to test the effect of different types of bedding material (straw vs. wood shavings vs. coarse bark mulch vs. cotton) on the prevalence of pododermatitis in 30 fattening Separator rabbits. Once a week, the four experimental runs were assessed for contamination (macroscopic contamination by faeces/urine/litter) as well as for lesion scoring of the footpad. In addition, the water and ammonia binding behaviour of bedding was tested. The most favourable values as to dry matter, NH3 (p < .05) and also scoring of the litter were generally observed when cotton was used as bedding material. This is probably due to the high water binding capacity, the faster water release and the structural effects of the material itself (softness). By contrast, the structure of the coarse bark mulch was not suitable for binding liquids (especially urine) and led to increased contamination of the barrels. These results suggest that rabbits with an increased tendency to sore soles should be kept on bedding materials with desirable physical properties (e.g. cotton litter).

Application of the Matrix Method for Calculating Internal Equilibrium Constants and Complex Formation Microconstants

This review is devoted to the matrix method of simulating complex formation in solution. The applicability of the method to the interpretation of spectrometric and calorimetric data for systems containing simultaneously a large number of complex forms is shown. The works where this method was used to calculate the internal equilibrium constants and cis/trans isomerization constants were analyzed. Using the matrix simulation method, the cooperativity of ammonia binding by Mg2+, Ca2+, Zn2+, Cd2+, and Ni2+ ions in aqueous solution was demonstrated. For halide complexes of Pt(II), Pd(II) and Au(III) with a square-planar structure, the difference between the mutual influence of ligands in the cis and trans isomers is shown. The question of verifying the hypothesis about the geometry of the coordination polyhedron of Cu2+ compounds formed in the processes of equilibrium stepwise complex formation in mixed solvents is considered.

Ammonia as a substrate-water analogue in photosynthetic water oxidation: Influence on activation barrier of the O2-formation step

Information on binding and rearrangement of pivotal water molecules could support understanding of light-driven water oxidation at the catalytic Mn4CaO5 cluster of photosystem II (PSII). To address this point, the binding of ammonia (NH3)—a possible substrate-water analogue—has been investigated and discussed in the context of putative reaction mechanisms. By time-resolved detection of O2 formation after light-flash excitation, we discriminate three NH3/NH4+ binding sites jointly characterized by a Km value around 25 mM (of NH4+), but differing in their influence on the O2-formation step. At 100 mM NH4Cl (pH 7.5), we observe (1) a PSII fraction with complete inhibition of O2-formation, (2) fast O2-formation with a time constant of 1.7 ms at 20 °C (Fast-PSII), and (3) slow O2-formation with a time constant of 36 ms at 20 °C (Slow-PSII). For the Fast-PSII, we determine an activation enthalpy of 223 ± 11 meV. Activation enthalpy and entropy of the Fast-PSII are essentially identical to the corresponding figures in the absence NH3/NH4+ binding. For the Slow-PSII, the activation enthalpy is 323 ± 11 meV and thus significantly increased, whereas the activation entropy remains essentially unchanged. We conclude: (1) The fully-inhibitory binding site could relate to bound NH3 replacing one of the two substrate-water molecules. (2) The Fast-PSII may relate to NH3/NH4+ binding in the S2-state of PSII followed by unbinding before onset of the OO bond formation step, but also more intricate mechanisms are not excluded. (3) In the Slow-PSII, NH3/NH4+ binding increases the energetic barrier of the OO bond formation step significantly.

Ab Initio Screening of Metal Catecholates for Adsorption of Toxic Pnictogen Hydride Gases

Density functional theory was used to study interactions between phosphine and 17 functional groups, revealing that only the functional groups containing metal ions bind phosphine strongly. This led to a detailed examination of the adsorption of the pnictogen hydride gases ammonia, phosphine, and arsine on 33 metal catecholates in order to examine periodic trends in the binding strength for these adsorbates. Phosphine and arsine adsorption is primarily driven by donation of electron density from the adsorbate molecule to the metal atom, while ammonia binding involves both electron donation and a significant Coulomb attraction between the negatively charged N atom and the positive metal atom. This Coulomb effect results in notably different binding behavior for ammonia than for phosphine and arsine, which are quite similar to each other. Generally, for metals on the left side of the periodic table, the Coulomb effect dominates in ammonia adsorption, and metals on the right side of the periodic table have l...

Inhibitory and Non-Inhibitory NH3 Binding at the Water-Oxidizing Manganese Complex of Photosystem II Suggests Possible Sites and a Rearrangement Mode of Substrate Water Molecules.

The identity and rearrangements of substrate water molecules in photosystem II (PSII) water oxidation are of great mechanistic interest and addressed herein by comprehensive analysis of NH4+/NH3 binding. Time-resolved detection of O2 formation and recombination fluorescence as well as Fourier transform infrared (FTIR) difference spectroscopy on plant PSII membrane particles reveals the following. (1) Partial inhibition in NH4Cl buffer occurs with a pH-independent binding constant of ∼25 mM, which does not result from decelerated O2 formation, but from complete blockage of a major PSII fraction (∼60%) after reaching the Mn(IV)4 (S3) state. (2) The non-inhibited PSII fraction advances through the reaction cycle, but modified nuclear rearrangements are suggested by FTIR difference spectroscopy. (3) Partial inhibition can be explained by anticooperative (mutually exclusive) NH3 binding to one inhibitory and one non-inhibitory site; these two sites may correspond to two water molecules terminally bound to the "dangling" Mn ion. (4) Unexpectedly strong modifications of the FTIR difference spectra suggest that in the non-inhibited PSII, ammonia binding obliterates the need for some of the nuclear rearrangements occurring in the S2-S3 transition as well as their reversal in the O2 formation transition, in line with the carousel mechanism [Askerka, M., et al. (2015) Biochemistry 54, 5783]. (5) We observe the same partial inhibition of PSII by NH4Cl also for thylakoid membranes prepared from mesophilic and thermophilic cyanobacteria, suggesting that the results described above are valid for plant and cyanobacterial PSII.

Molecular characterization of two Rhesus glycoproteins from the euryhaline freshwater white-rimmed stingray, Himantura signifer, and changes in their transcript levels and protein abundance in the gills, kidney, and liver during brackish water acclimation.

Himantura signifer is a freshwater stingray which inhabits rivers in Southeast Asia. It is ammonotelic in fresh water, but retains the capacities of urea synthesis and ureosmotic osmoregulation to survive in brackish water. This study aimed to elucidate the roles of Rhesus glycoproteins (Rhgp), which are known to transport ammonia, in conserving nitrogen (N) in H. signifer during brackish water acclimation when N became limited resulting from increased hepatic urea synthesis. The complete coding sequence of rhbg from H. signifer consisted of 1383bp, encoding 460 amino acids with an estimated molecular mass of 50.5kDa, while that of rhcg comprised 1395bp, encoding for 464 amino acids with an estimated molecular mass of 50.8kDa. The deduced amino sequences of Rhbg and Rhcg contained ammonia binding sites, which could recruit NH4+ to be deprotonated, and a hydrophobic pore with two histidine residues, which could mediate the transport of NH3. Our results indicated for the first time that brackish water acclimation resulted in significant decreases in the expression levels of rhbg/Rhbg and rhcg/Rhcg in the gills of H. signifer, which offered a mechanistic explanation of brackish water-related decreased ammonia excretion reported elsewhere. Furthermore, rhbg/Rhbg expression levels increased significantly in the liver of H. signifer during brackish water acclimation, indicating that the ammonia produced by extra-hepatic tissues and released into the blood could be channeled into the liver for increased urea synthesis. Overall, these results lend support to the proposition that H. signifer becomes N-limited upon utilizing urea as an osmolyte in brackish water.

Characterization of ammonia binding to the second coordination shell of the oxygen-evolving complex of photosystem II.

The second-shell ammonia binding sites near the OEC (oxygen-evolving complex) of PSII are characterized by combined Continuum Electrostatic/Monte Carlo (MCCE), QM/MM and DFT calculations and compared with new and earlier experimental measurements. MCCE shows ammonia has significant affinity at 6 positions but only two significantly influence the OEC. Although the pKa of ammonium ion is 9.25, it is calculated to only bind as NH3, in agreement with its low affinity at low pH. The calculations also help explain the experimentally observed competitive binding of ammonia with chloride. Ammonia and Cl- compete for one site. Electrostatic interactions cause Cl- to effect ammonia at two other sites. Cl- stabilizes the multiline g = 2.0 form of the S2 state (OEC Mn oxidation state 3444) while ammonia only binds in the g = 4.1 form of the S2 state (oxidation state 4443) due to the movement of the positive charge between Mn1 and Mn4. One ammonia binds near Mn4 and shares a proton with D2-K317, making the ion pair NH4+K3170D61-, making ammonia binding sensitive to the K317A mutation. The affinity of ammonia is also dependent on the protonation state of water 2, a primary ligand to Mn4.

有機・無機複合薄膜を修飾した水晶振動子センサによるアンモニアおよびホルムアルデヒドの二成分ガス検知

Quartz crystal microbalance (QCM) gas sensors based on the alternate deposition of TiO2 and polymers were developed for the detection of ammonia and formaldehyde odors. For sensitive detection of both analytes, poly(acrylic acid), PAA, and poly(allyamine hydrochloride), PAH, were employed in the film along with TiO2 thin layers that were prepared by the gas-phase surface sol-gel (GSSG) process. The alternate (TiO2/PAA/TiO2/PAH) film-deposited QCM sensor showed linear responses to ammonia in the concentration range 0.5-40 ppm and to formaldehyde in the concentration range 0.3-20 ppm. The sensor response was very fast, stable, and repeatable. The ammonia or formaldehyde binding is based on the acid-base interaction to the free carboxyl or amino groups of PAA or PAH. TiO2 thin layers deposited by the GSSG process played significant roles in preventing the electrostatic interaction of polymer layers and in improving the diffusion of gaseous analytes inside the film.

The O2-Evolving Complex of Photosystem II: Recent Insights from Quantum Mechanics/Molecular Mechanics (QM/MM), Extended X-ray Absorption Fine Structure (EXAFS), and Femtosecond X-ray Crystallography Data.

Efficient photoelectrochemical water oxidation may open a way to produce energy from renewable solar power. In biology, generation of fuel due to water oxidation happens efficiently on an immense scale during the light reactions of photosynthesis. To oxidize water, photosynthetic organisms have evolved a highly conserved protein complex, Photosystem II. Within that complex, water oxidation happens at the CaMn4O5 inorganic catalytic cluster, the so-called oxygen-evolving complex (OEC), which cycles through storage "S" states as it accumulates oxidizing equivalents and produces molecular oxygen. In recent years, there has been significant progress in understanding the OEC as it evolves through the catalytic cycle. Studies have combined conventional and femtosecond X-ray crystallography with extended X-ray absorption fine structure (EXAFS) and quantum mechanics/molecular mechanics (QM/MM) methods and have addressed changes in protonation states of μ-oxo bridges and the coordination of substrate water through the analysis of ammonia binding as a chemical analog of water. These advances are thought to be critical to understanding the catalytic cycle since protonation states regulate the relative stability of different redox states and the geometry of the OEC. Therefore, establishing the mechanism for substrate water binding and the nature of protonation/redox state transitions in the OEC is essential for understanding the catalytic cycle of O2 evolution. The structure of the dark-stable S1 state has been a target for X-ray crystallography for the past 15 years. However, traditional X-ray crystallography has been hampered by radiation-induced reduction of the OEC. Very recently, a revolutionary X-ray free electron laser (XFEL) technique was applied to PSII to reveal atomic positions at 1.95 Å without radiation damage, which brought us closer than ever to establishing the ultimate structure of the OEC in the S1 state. However, the atom positions in this crystal structure are still not consistent with high-resolution EXAFS spectroscopy, partially due to the poorly resolved oxygen positions next to Mn centers and partial reduction due to extended dark adaptation of the sample. These inconsistencies led to the new models of the OEC with an alternative low oxidation state and raised questions on the protonation state of the cluster, especially the O5 μ-oxo bridge. This Account summarizes the most recent models of the OEC that emerged from QM/MM, EXAFS and femtosecond X-ray crystallography methods. When PSII in the S1 state is exposed to light, the S1 state is advanced to the higher oxidation states and eventually binds substrate water molecules. Identifying the substrate waters is of paramount importance for establishing the water-oxidation mechanism but is complicated by a large number of spectroscopically similar waters. Water analogues can, therefore, be helpful because they serve as spectroscopic markers that help to track the motion of the substrate waters. Due to a close structural and electronic similarity to water, ammonia has been of particular interest. We review three competing hypotheses on substrate water/ammonia binding and compile theoretical and experimental evidence to support them. Binding of ammonia as a sixth ligand to Mn4 during the S1 → S2 transition seems to satisfy most of the criteria, especially the most compelling recent EPR data on D1-D61A mutated PSII. Such a binding mode suggests delivery of water from the "narrow" channel through a "carousel" rearrangement of waters around Mn4 upon the S2 → S3 transition. An alternative hypothesis suggests water delivery through the "large" channel on the Ca side. However, both water delivery paths lead to a similar S3 structure, seemingly reaching consensus on the nature of the last detectable S-state intermediate in the Kok cycle before O2 evolution.

Ammonia Binding in the Second Coordination Sphere of the Oxygen-Evolving Complex of Photosystem II.

Ammonia binds to two sites in the oxygen-evolving complex (OEC) of Photosystem II (PSII). The first is as a terminal ligand to Mn in the S2 state, and the second is at a site outside the OEC that is competitive with chloride. Binding of ammonia in this latter secondary site results in the S2 state S = (5)/2 spin isomer being favored over the S = (1)/2 spin isomer. Using electron paramagnetic resonance spectroscopy, we find that ammonia binds to the secondary site in wild-type Synechocystis sp. PCC 6803 PSII, but not in D2-K317A mutated PSII that does not bind chloride. By combining these results with quantum mechanics/molecular mechanics calculations, we propose that ammonia binds in the secondary site in competition with D1-D61 as a hydrogen bond acceptor to the OEC terminal water ligand, W1. Implications for the mechanism of ammonia binding via its primary site directly to Mn4 in the OEC are discussed.

S3 State of the O2-Evolving Complex of Photosystem II: Insights from QM/MM, EXAFS, and Femtosecond X-ray Diffraction

The oxygen-evolving complex (OEC) of photosystem II has been studied in the S3 state by electron paramagnetic resonance, extended X-ray absorption fine structure (EXAFS), and femtosecond X-ray diffraction (XRD). However, the actual structure of the OEC in the S3 state has yet to be established. Here, we apply hybrid quantum mechanics/molecular mechanics methods and propose a structural model that is consistent with EXAFS and XRD. The model supports binding of water ligands to the cluster in the S2 → S3 transition through a carousel rearrangement around Mn4, inspired by studies of ammonia binding.