| 1. |
- Guo, Y., et al.
(författare)
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Reversible Structural Isomerization of Nature's Water Oxidation Catalyst Prior to O-O Bond Formation
- 2022
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 144:26, s. 11736-11747
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Tidskriftsartikel (refereegranskat)abstract
- Photosynthetic water oxidation is catalyzed by a manganese-calcium oxide cluster, which experiences five "S-states" during a light-driven reaction cycle. The unique "distorted chair"-like geometry of the Mn4CaO5(6)cluster shows structural flexibility that has been frequently proposed to involve "open" and "closed"-cubane forms from the S1 to S3states. The isomers are interconvertible in the S1 and S2states, while in the S3state, the open-cubane structure is observed to dominate inThermosynechococcus elongatus (cyanobacteria) samples. In this work, using density functional theory calculations, we go beyond the S3+Yzstate to the S3nYz•→ S4+Yzstep, and report for the first time that the reversible isomerism, which is suppressed in the S3+Yzstate, is fully recovered in the ensuing S3nYz•state due to the proton release from a manganese-bound water ligand. The altered coordination strength of the manganese-ligand facilitates formation of the closed-cubane form, in a dynamic equilibrium with the open-cubane form. This tautomerism immediately preceding dioxygen formation may constitute the rate limiting step for O2formation, and exert a significant influence on the water oxidation mechanism in photosystem II.
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| 2. |
- Simon, Philipp S., et al.
(författare)
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Capturing the sequence of events during the water oxidation reaction in photosynthesis using XFELs
- 2023
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Ingår i: FEBS Letters. - : John Wiley & Sons. - 0014-5793 .- 1873-3468. ; 597:1, s. 30-37
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Tidskriftsartikel (refereegranskat)abstract
- Ever since the discovery that Mn was required for oxygen evolution in plants by Pirson in 1937 and the period-four oscillation in flash-induced oxygen evolution by Joliot and Kok in the 1970s, understanding of this process has advanced enormously using state-of-the-art methods. The most recent in this series of innovative techniques was the introduction of X-ray free-electron lasers (XFELs) a decade ago, which led to another quantum leap in the understanding in this field, by enabling operando X-ray structural and X-ray spectroscopy studies at room temperature. This review summarizes the current understanding of the structure of Photosystem II (PS II) and its catalytic centre, the Mn4CaO5 complex, in the intermediate Si (i = 0–4)-states of the Kok cycle, obtained using XFELs.
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| 3. |
- Fuller, Franklin D, et al.
(författare)
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Drop-on-demand sample delivery for studying biocatalysts in action at X-ray free-electron lasers
- 2017
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Ingår i: Nature Methods. - : Macmillan Publishers Ltd.. - 1548-7091 .- 1548-7105. ; 14, s. 443-449
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Tidskriftsartikel (refereegranskat)abstract
- X-ray crystallography at X-ray free-electron laser sources is a powerful method for studying macromolecules at biologically relevant temperatures. Moreover, when combined with complementary techniques like X-ray emission spectroscopy, both global structures and chemical properties of metalloenzymes can be obtained concurrently, providing insights into the interplay between the protein structure and dynamics and the chemistry at an active site. The implementation of such a multimodal approach can be compromised by conflicting requirements to optimize each individual method. In particular, the method used for sample delivery greatly affects the data quality. We present here a robust way of delivering controlled sample amounts on demand using acoustic droplet ejection coupled with a conveyor belt drive that is optimized for crystallography and spectroscopy measurements of photochemical and chemical reactions over a wide range of time scales. Studies with photosystem II, the phytochrome photoreceptor, and ribonucleotide reductase R2 illustrate the power and versatility of this method.
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| 4. |
- Messinger, Johannes, et al.
(författare)
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Photosynthesis : from natural to artificial
- 2014
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Ingår i: Physical Chemistry, Chemical Physics - PCCP. - : Royal Society of Chemistry (RSC). - 1463-9076 .- 1463-9084. ; 16:24, s. 11810-11811
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
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| 5. |
- Sierra, Raymond G., et al.
(författare)
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Nanoflow electrospinning serial femtosecond crystallography
- 2012
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Ingår i: Acta Crystallographica Section D. - : Wiley-Blackwell. - 0907-4449 .- 1399-0047. ; 68, s. 1584-1587
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Tidskriftsartikel (refereegranskat)abstract
- An electrospun liquid microjet has been developed that delivers protein microcrystal suspensions at flow rates of 0.14-3.1 mu l min(-1) to perform serial femtosecond crystallography (SFX) studies with X-ray lasers. Thermolysin microcrystals flowed at 0.17 mu l min(-1) and diffracted to beyond 4 angstrom resolution, producing 14 000 indexable diffraction patterns, or four per second, from 140 mu g of protein. Nanoflow electrospinning extends SFX to biological samples that necessitate minimal sample consumption.
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| 6. |
- Cheah, Mun Hon, et al.
(författare)
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Assessment of the manganese cluster’s oxidation state via photoactivation of photosystem II microcrystals
- 2020
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 117:1, s. 141-145
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Tidskriftsartikel (refereegranskat)abstract
- Knowledge of the manganese oxidation states of the oxygen-evolving Mn4CaO5 cluster in photosystem II (PSII) is crucial toward understanding the mechanism of biological water oxidation. There is a 4 decade long debate on this topic that historically originates from the observation of a multiline electron paramagnetic resonance (EPR) signal with effective total spin of S = 1/2 in the singly oxidized S2 state of this cluster. This signal implies an overall oxidation state of either Mn(III)3Mn(IV) or Mn(III)Mn(IV)3 for the S2 state. These 2 competing assignments are commonly known as “low oxidation (LO)” and “high oxidation (HO)” models of the Mn4CaO5 cluster. Recent advanced EPR and Mn K-edge X-ray spectroscopy studies converge upon the HO model. However, doubts about these assignments have been voiced, fueled especially by studies counting the number of flash-driven electron removals required for the assembly of an active Mn4CaO5 cluster starting from Mn(II) and Mn-free PSII. This process, known as photoactivation, appeared to support the LO model since the first oxygen is reported to evolve already after 7 flashes. In this study, we improved the quantum yield and sensitivity of the photoactivation experiment by employing PSII microcrystals that retained all protein subunits after complete manganese removal and by oxygen detection via a custom built thin-layer cell connected to a membrane inlet mass spectrometer. We demonstrate that 9 flashes by a nanosecond laser are required for the production of the first oxygen, which proves that the HO model provides the correct description of the Mn4CaO5 cluster’s oxidation states.
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| 7. |
- de Lichtenberg, Casper, 1989-
(författare)
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Time-resolved Structural and Mechanistic Studies of Water Oxidation in Photosystem II : water here, water there, water everywhere
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Oxygenic photosynthesis is undisputedly one of the most important chemical processes for human life on earth as it not only fills the atmosphere with the oxygen that we need to breathe, but also sustains the accumulation of biomass, which is not only used as nourishment but is also present in almost every aspect of our lives as building material, textiles in clothes and furniture, or even as living decorations to name a few.The photosynthetic water-splitting mechanism is catalyzed by a water:plastoquinone oxido-reductase by the name of photosystem II (PSII), which is embedded in the thylakoid membranes of plants, algae and cyanobacteria. As it is excited by light, charge separation occurs in the reaction center of the protein and an electron is extracted by oxidation of Mn4Ca-cluster, that constitutes the active site for the water splitting reaction in PSII. When the Mn4Ca-cluster has been oxidized 4 times, it forms an oxygen-oxygen bond between two water derived ligands bound to the Mn4Ca-cluster and returns to the lowest oxidation state of the catalytic cycle. Understanding what ligands of the cluster that are used in the water splitting reaction is the key to unlocking the underlying chemical mechanism.In this thesis I describe investigations, with room temperature X-ray diffraction (XRD) and X-ray emission spectroscopy (XES) on PSII microcrystals, of how the active site looks in all the stable intermediate oxidation states. Furthermore I describe how we uncovered the sequence of events that lead to insertion of an additional water ligand in the S2-S3 state transition of the catalytic cycle.Furthermore, through time-resolved membrane-inlet mass spectrometry (TR-MIMS) measurements of the isotopic equilibration of the substrate waters with the bulk in conditions that induce different electron magnetic resonance (EPR) spectroscopic signatures, I present evidence that the exchange of the slowly exchanging substrate water Ws is controlled by a dynamic equilibrium between conformations in the S2-state that give rise to either the low-spin multiline (LS-ML) signal or the high-spin (HS) signal. Based on the crystal structures and litterature suggestions for the conformation of the HS state different scenarios were presented for the assignment of Ws and how it exchanges. This analysis is discussed in the context of all semi-stable intermediate oxidation states in the Kok cycle.To further the understanding of this equilibrium, I also studied a selection of mutants positioned at strategic places in the vicinity of the different proposed substrates and at points that were suggested to be critical for substrate entry. With the combination of TR-MIMS and EPR, I reached the conclusion that by mutating valine 185 to asparagine, the water bound A-type conformation was stabilized, meanwhile in the mutant where aspartate 61 was mutated to alanine I observed that the barrier of the equilibrium between the exchanging conformations was so high that the interchange between them was arrested at room temperature. Additionally the retardation of the substrate exchange rates in the S3-states fit best with D61 being in the vicinity of the fast exchanging water. With this information we found the data best explained in a scenario where the water insertion of the S2-S3 transition was determining the if O-O bond formation occurred between the waters that were W2 and W3 or W2 and O5 in the S2 state. In addition, by mutation of glutamate 189 to glutamine that this residue is not important for the exchange of substrate waters in the S2 or the S3 states.Finally I use a combination of substrate labelling with TR-MIMS and time resolved labelling of the waters that ligate the Mn4Ca-cluster to show that the briding oxygen O5 is exchanging with a near identical rate to Ws, further supporting the assignment that Ws=O5.In conclusion, O-O bond formation most likely occurs between W2 (Wf) and O5 (Ws) via an oxo-oxyl radical coupling mechanism. The newly inserted water thus represents the slow exchanging water of the following S-state cycle.
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| 8. |
- Han, Guangye, et al.
(författare)
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Molecular basis for turnover inefficiencies (misses) during water oxidation in photosystem II
- 2022
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Ingår i: Chemical Science. - : Royal Society of Chemistry. - 2041-6520 .- 2041-6539. ; 13:29, s. 8667-8678
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Tidskriftsartikel (refereegranskat)abstract
- Photosynthesis stores solar light as chemical energy and efficiency of this process isv highly important. The electrons required for CO2 reduction are extracted from water in a reaction driven by light-induced charge separations in the Photosystem II reaction center and catalyzed by the CaMn4O5-cluster. This cyclic process involves five redox intermediates known as the S-0-S-4 states. In this study, we quantify the flash-induced turnover efficiency of each S state by electron paramagnetic resonance spectroscopy. Measurements were performed in photosystem II membrane preparations from spinach in the presence of an exogenous electron acceptor at selected temperatures between -10 degrees C and +20 degrees C and at flash frequencies of 1.25, 5 and 10 Hz. The results show that at optimal conditions the turnover efficiencies are limited by reactions occurring in the water oxidizing complex, allowing the extraction of their S state dependence and correlating low efficiencies to structural changes and chemical events during the reaction cycle. At temperatures 10 degrees C and below, the highest efficiency (i.e. lowest miss parameter) was found for the S-1 -> S-2 transition, while the S-2 -> S-3 transition was least efficient (highest miss parameter) over the whole temperature range. These electron paramagnetic resonance results were confirmed by measurements of flash-induced oxygen release patterns in thylakoid membranes and are explained on the basis of S state dependent structural changes at the CaMn4O5-cluster that were determined recently by femtosecond X-ray crystallography. Thereby, possible "molecular errors" connected to the e(-) transfer, H+ transfer, H2O binding and O-2 release are identified.
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| 9. |
- Koroidov, Sergey, et al.
(författare)
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Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation
- 2014
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 11:17, s. 6299-6304
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Tidskriftsartikel (refereegranskat)abstract
- Cyanobacteria, algae and plants oxidize water to the O2 we breathe, and consume CO2 during the synthesis of biomass. Although these vital processes are functionally and structurally well separated in photosynthetic organisms, there is a long-debated role for CO2/HCO3− in water oxidation. Using membrane-inlet mass spectrometry we demonstrate that HCO3− acts as a mobile proton acceptor that helps to transport the protons produced inside of photosystem II by water-oxidation out into the chloroplast's lumen, resulting in a light-driven production of O2 and CO2. Depletion of HCO3− from the media leads, in the absence of added buffers, to a reversible down-regulation of O2 production by about 20%. These findings add a previously unidentified component to the regulatory network of oxygenic photosynthesis, and conclude the more than 50-y-long quest for the function of CO2/ HCO3− in photosynthetic water oxidation.
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| 10. |
- Shevela, Dmitriy, 1979-, et al.
(författare)
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'Birth defects' of photosystem II make it highly susceptible to photodamage during chloroplast biogenesis
- 2019
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Ingår i: Physiologia Plantarum. - : Wiley-Blackwell. - 0031-9317 .- 1399-3054. ; 166:1, s. 165-180
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Tidskriftsartikel (refereegranskat)abstract
- High solar flux is known to diminish photosynthetic growth rates, reducing biomass productivity and lowering disease tolerance. Photosystem II (PSII) of plants is susceptible to photodamage (also known as photoinactivation) in strong light, resulting in severe loss of water oxidation capacity and destruction of the water‐oxidizing complex (WOC). The repair of damaged PSIIs comes at a high energy cost and requires de novo biosynthesis of damaged PSII subunits, reassembly of the WOC inorganic cofactors and membrane remodeling. Employing membrane‐inlet mass spectrometry and O2‐polarography under flashing light conditions, we demonstrate that newly synthesized PSII complexes are far more susceptible to photodamage than are mature PSII complexes. We examined these ‘PSII birth defects’ in barley seedlings and plastids (etiochloroplasts and chloroplasts) isolated at various times during de‐etiolation as chloroplast development begins and matures in synchronization with thylakoid membrane biogenesis and grana membrane formation. We show that the degree of PSII photodamage decreases simultaneously with biogenesis of the PSII turnover efficiency measured by O2‐polarography, and with grana membrane stacking, as determined by electron microscopy. Our data from fluorescence, QB‐inhibitor binding, and thermoluminescence studies indicate that the decline of the high‐light susceptibility of PSII to photodamage is coincident with appearance of electron transfer capability QA− → QB during de‐etiolation. This rate depends in turn on the downstream clearing of electrons upon buildup of the complete linear electron transfer chain and the formation of stacked grana membranes capable of longer‐range energy transfer.
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