Mutation-induced shift of the photosystem II active site reveals insight into conserved water channels.

Photosystem II (PSII) is the water-plastoquinone photo-oxidoreductase central to oxygenic photosynthesis. PSII has been extensively studied for its ability to catalyze light-driven water oxidation at a MnCaO cluster called the oxygen-evolving complex (OEC). Despite these efforts, the complete reaction mechanism for water oxidation by PSII is still heavily debated.

Independent Mutation of Two Bridging Carboxylate Ligands Stabilizes Alternate Conformers of the Photosynthetic O-Evolving MnCaO Cluster in Photosystem II.

The O-evolving MnCaO cluster in photosystem II is ligated by six carboxylate residues. One of these is D170 of the D1 subunit. This carboxylate bridges between one Mn ion (Mn4) and the Ca ion.

Mutation of a metal ligand stabilizes the high-spin form of the S state in the O-producing MnCaO cluster of photosystem II.

The residue D1-D170 bridges Mn4 with the Ca ion in the O-evolving MnCaO cluster of Photosystem II. Recently, the D1-D170E mutation was shown to substantially alter the S-minus-S FTIR difference spectra [Debus RJ (2021) Biochemistry 60:3841-3855]. The mutation was proposed to alter the equilibrium between different Jahn-Teller conformers of the S state such that (i) a different S state conformer is stabilized in D1-D170E than in wild-type and (ii) the S to S transition in D1-D170E produces a high-spin form of the S state rather than the low-spin form that is produced in wild-type.

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

Photosystem II (PSII) enables global-scale, light-driven water oxidation. Genetic manipulation of PSII from the mesophilic cyanobacterium 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.

Alteration of the O-Producing MnCa Cluster in Photosystem II by the Mutation of a Metal Ligand.

The O-evolving MnCa cluster in photosystem II (PSII) is arranged as a distorted MnCa cube that is linked to a fourth Mn ion (denoted as Mn4) by two oxo bridges. The Mn4 and Ca ions are bridged by residue D1-D170. This is also the only residue known to participate in the high-affinity Mn(II) site that participates in the light-driven assembly of the MnCa cluster.

Structure of a monomeric photosystem II core complex from a cyanobacterium acclimated to far-red light reveals the functions of chlorophylls d and f.

Far-red light (FRL) photoacclimation in cyanobacteria provides a selective growth advantage for some terrestrial cyanobacteria by expanding the range of photosynthetically active radiation to include far-red/near-infrared light (700-800 nm). During this photoacclimation process, photosystem II (PSII), the water:plastoquinone photooxidoreductase involved in oxygenic photosynthesis, is modified. The resulting FRL-PSII is comprised of FRL-specific core subunits and binds chlorophyll (Chl) d and Chl f molecules in place of several of the Chl a molecules found when cells are grown in visible light.

The exchange of the fast substrate water in the S state of photosystem II is limited by diffusion of bulk water through channels - implications for the water oxidation mechanism.

The molecular oxygen we breathe is produced from water-derived oxygen species bound to the MnCaO cluster in photosystem II (PSII). Present research points to the central oxo-bridge O5 as the 'slow exchanging substrate water (W)', while, in the S state, the terminal water ligands W2 and W3 are both discussed as the 'fast exchanging substrate water (W)'. A critical point for the assignment of W is whether or not its exchange with bulk water is limited by barriers in the channels leading to the MnCaO cluster.

Pulse EPR Spectroscopic Characterization of the S State of the Oxygen-Evolving Complex of Photosystem II Isolated from .

The S state is the last semi-stable state in the water splitting reaction that is catalyzed by the MnOCa cluster that makes up the oxygen-evolving complex (OEC) of photosystem II (PSII). Recent high-field/frequency (95 GHz) electron paramagnetic resonance (EPR) studies of PSII isolated from the thermophilic cyanobacterium have found broadened signals induced by chemical modification of the S state. These signals are ascribed to an S form that contains a five-coordinate Mn center bridged to a cuboidal MnOCa unit.

Roles of D1-Glu189 and D1-Glu329 in O Formation by the Water-Splitting MnCa Cluster in Photosystem II.

During the catalytic step that precedes O-O bond formation in Photosystem II, a water molecule deprotonates and moves next to the water-splitting MnCa cluster's O5 oxo bridge. The relocated oxygen, known as O6 or O, may serve as a substrate, combining with O5 to form O during the final step in the catalytic cycle, or may be positioned to become a substrate during the next catalytic cycle. Recent serial femtosecond X-ray crystallographic studies show that the flexibility of D1-E189 plays a critical role in facilitating the relocation of O/O.

D1-S169A substitution of photosystem II reveals a novel S-state structure.

In photosystem II (PSII), photosynthetic water oxidation occurs at the O-evolving complex (OEC), a tetramanganese-calcium cluster that cycles through light-induced redox intermediates (S-S) to produce oxygen from two substrate water molecules. The OEC is surrounded by a hydrogen-bonded network of amino-acid residues that plays a crucial role in proton transfer and substrate water delivery. Previously, we found that D1-S169 was crucial for water oxidation and its mutation to alanine perturbed the hydrogen-bonding network.

Determining the Electronic Structure of Paramagnetic Intermediates in membrane proteins: A high-resolution 2D H hyperfine sublevel correlation study of the redox-active tyrosines of photosystem II.

The photosynthetic reaction center, photosystem II (PSII), catalyzes one of the most energetically demanding reactions in nature by using light energy to drive water oxidation. The four-electron water oxidation reaction occurs at the tetranuclear manganese‑calcium-oxo (MnCa-oxo) cluster that is present in the oxygen-evolving complex (OEC) of PSII. The water oxidation reaction is facilitated by proton-coupled electron transfer (PCET) at the redox-active tyrosine residue, Y, in the OEC which is one of the two symmetric tyrosine residues, Y and Y, in PSII.

Insights into Proton-Transfer Pathways during Water Oxidation in Photosystem II.

Water oxidation by photosystem II (PSII) involves the release of O, electrons, and protons at the oxygen-evolving complex (OEC). These processes are facilitated by a hydrogen-bonded network of amino acid residues and waters surrounding the OEC. It is crucial to probe the proton-transfer pathways from the OEC as proton release helps to maintain the charge balance required for efficient water oxidation.

One of the Substrate Waters for O Formation in Photosystem II Is Provided by the Water-Splitting MnCaO Cluster's Ca Ion.

During the catalytic step immediately prior to O-O bond formation in Photosystem II, a water molecule deprotonates and moves next to the water-splitting MnCaO cluster's O5 oxo bridge. Considerable evidence identifies O5 as one of the two substrate waters that ultimately form O. The relocated oxygen, known as O6 or O, may be the second.

Bicarbonate rescues damaged proton-transfer pathway in photosystem II.

The membrane-protein complex photosystem II (PSII) catalyzes photosynthetic water oxidation. Proton transfer plays an integral role in the catalytic cycle of water oxidation by maintaining charge balance to regulate and ensure the efficiency of the process. The hydrogen-bonded amino-acid residues that surround the oxygen-evolving complex (OEC) provide an efficient pathway for proton removal.

D1-S169A Substitution of Photosystem II Perturbs Water Oxidation.

In photosystem II (PSII), photosynthetic water oxidation occurs at the tetramanganese-calcium cluster that cycles through light-induced intermediates (S-S) to produce oxygen from two substrate waters. The surrounding hydrogen-bonded amino acid residues and waters form channels that facilitate proton transfer and substrate water delivery, thereby ensuring efficient water oxidation. The residue D1-S169 lies in the "narrow" channel and forms hydrogen bonds with the MnCaO cluster via waters W1 and Wx.

Impact of D1-V185 on the Water Molecules That Facilitate O Formation by the Catalytic MnCaO Cluster in Photosystem II.

The oxidations of the O-evolving MnCaO cluster in Photosystem II are coupled to the release of protons to the thylakoid lumen via one or more proton egress pathways. These pathways are comprised of extensive networks of hydrogen-bonded water molecules and amino acid side chains. The hydrophobic residue, D1-V185, is adjacent to numerous water molecules in one of these pathways.

Structural Effects of Ammonia Binding to the MnCaO Cluster of Photosystem II.

The MnCaO oxygen-evolving complex (OEC) of photosystem II catalyzes the light-driven oxidation of two substrate waters to molecular oxygen. ELDOR-detected NMR along with computational studies indicated that ammonia, a substrate analogue, binds as a terminal ligand to the Mn4A ion trans to the O5 μ oxido bridge. Results from electron spin echo envelope modulation (ESEEM) spectroscopy confirmed this and showed that ammonia hydrogen bonds to the carboxylate side chain of D1-Asp61.

Substitution of the D1-Asn site in photosystem II of cyanobacteria mimics the chloride-binding characteristics of spinach photosystem II.

Photoinduced water oxidation at the O-evolving complex (OEC) of photosystem II (PSII) is a complex process involving a tetramanganese-calcium cluster that is surrounded by a hydrogen-bonded network of water molecules, chloride ions, and amino acid residues. Although the structure of the OEC has remained conserved over eons of evolution, significant differences in the chloride-binding characteristics exist between cyanobacteria and higher plants. An analysis of amino acid residues in and around the OEC has identified residue 87 in the D1 subunit as the only significant difference between PSII in cyanobacteria and higher plants.

Evidence from FTIR Difference Spectroscopy That a Substrate HO Molecule for O Formation in Photosystem II Is Provided by the Ca Ion of the Catalytic MnCaO Cluster.

The O-producing MnCaO catalyst in photosystem II oxidizes two water molecules (substrate) to produce one O molecule. Considerable evidence supports the identification of one of the two substrate waters as the MnCaO cluster's oxo bridge known as O. The identity of the second substrate water molecule is less clear.

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.

Ammonia Binds to the Dangler Manganese of the Photosystem II Oxygen-Evolving Complex.

High-resolution X-ray structures of photosystem II reveal several potential substrate binding sites at the water-oxidizing/oxygen-evolving 4MnCa cluster. Aspartate-61 of the D1 protein hydrogen bonds with one such water (W1), which is bound to the dangler Mn4A of the oxygen-evolving complex. Comparison of pulse EPR spectra of (14)NH3 and (15)NH3 bound to wild-type Synechocystis PSII and a D1-D61A mutant lacking this hydrogen-bonding interaction demonstrates that ammonia binds as a terminal NH3 at this dangler Mn4A site and not as a partially deprotonated bridge between two metal centers.

Probing the effect of mutations of asparagine 181 in the D1 subunit of photosystem II.

Efficient proton removal from the oxygen-evolving complex (OEC) of photosystem II (PSII) and activation of substrate water molecules are some of the key aspects optimized in the OEC for high turnover rates. The hydrogen-bonding network around the OEC is critical for efficient proton transfer and for tuning the position and pKa values of the substrate water/hydroxo/oxo molecules. The D1-N181 residue is part of the hydrogen-bonding network on the active face of the OEC.

Warwick Hillier: a tribute.

Warwick Hillier (October 18, 1967-January 10, 2014) made seminal contributions to our understanding of photosynthetic water oxidation employing membrane inlet mass spectrometry and FTIR spectroscopy. This article offers a collection of historical perspectives on the scientific impact of Warwick Hillier's work and tributes to the personal impact his life and ideas had on his collaborators and colleagues.

FTIR studies of metal ligands, networks of hydrogen bonds, and water molecules near the active site Mn₄CaO₅ cluster in Photosystem II.

The photosynthetic conversion of water to molecular oxygen is catalyzed by the Mn₄CaO₅ cluster in Photosystem II and provides nearly our entire supply of atmospheric oxygen. The Mn₄CaO₅ cluster accumulates oxidizing equivalents in response to light-driven photochemical events within Photosystem II and then oxidizes two molecules of water to oxygen. The Mn₄CaO₅ cluster converts water to oxygen much more efficiently than any synthetic catalyst because its protein environment carefully controls the cluster's reactivity at each step in its catalytic cycle.