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Gergely Katona - One of the best experts on this subject based on the ideXlab platform.
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Asymmetry in serial femtosecond crystallography data
Acta Crystallographica Section A Foundations and Advances, 2017Co-Authors: Amit Sharma, Weixiao Yuan Wahlgren, Linda C. Johansson, Richard Neutze, Elin Dunevall, Gergely KatonaAbstract:Serial crystallography is an increasingly important approach to protein crystallography that exploits both X-ray free-electron laser (XFEL) and synchrotron radiation. Serial crystallography recovers complete X-ray diffraction data by processing and merging diffraction images from thousands of randomly oriented non-uniform microcrystals, of which all observations are partial Bragg reflections. Random fluctuations in the XFEL pulse energy spectrum, variations in the size and shape of microcrystals, integrating over millions of weak partial observations and instabilities in the XFEL beam position lead to new types of experimental errors. The quality of Bragg intensity estimates deriving from serial crystallography is therefore contingent upon assumptions made while modeling these data. Here it is observed that serial femtosecond crystallography (SFX) Bragg reflections do not follow a unimodal Gaussian distribution and it is recommended that an idealized assumption of single Gaussian peak profiles be relaxed to incorporate apparent asymmetries when processing SFX data. The phenomenon is illustrated by re-analyzing data collected from microcrystals of the Blastochloris viridis photosynthetic reaction center and comparing these intensity observations with conventional synchrotron data. The results show that skewness in the SFX observations captures the essence of the Wilson plot and an empirical treatment is suggested that can help to separate the diffraction Bragg intensity from the background.
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography
2016Co-Authors: Linda C. Johansson, Gergely Katona, David Arnlund, Thomas A. White, Daniel P. Deponte, Robert L. Shoeman, Anton Barty, Cecilia Wickstr, Amit Sharma, Garth J. WilliamsAbstract:Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8Å resolution and determine its serial femtosecond crystallography structure to 3.5Å resolution. Although every microcrystal is exposed to a dose of 33MGy, no signs of X-ray-induced radiation damage are visible in this integra
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography
Nature communications, 2013Co-Authors: Linda C. Johansson, Gergely Katona, David Arnlund, Cecilia Wickstrand, Thomas A. White, Daniel P. Deponte, Robert L. Shoeman, Anton Barty, Amit Sharma, Garth J. WilliamsAbstract:Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8 A resolution and determine its serial femtosecond crystallography structure to 3.5 A resolution. Although every microcrystal is exposed to a dose of 33 MGy, no signs of X-ray-induced radiation damage are visible in this integral membrane protein structure.
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography
Nature Communications, 2013Co-Authors: Linda C. Johansson, Gergely Katona, David Arnlund, Cecilia Wickstrand, Thomas A. White, Daniel P. Deponte, Robert L. Shoeman, Anton Barty, Amit Sharma, Garth J. WilliamsAbstract:Serial femtosecond crystallography is an X-ray free-electron-laser-based method that uses X-ray bursts to determine protein structures. Here the authors present the structure of a photosynthetic reaction centre, an integral membrane protein, achieved with no sign of X-ray-induced radiation damage. Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8 Å resolution and determine its serial femtosecond crystallography structure to 3.5 Å resolution. Although every microcrystal is exposed to a dose of 33 MGy, no signs of X-ray-induced radiation damage are visible in this integral membrane protein structure.
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Structural Characterization of Bacterioferritin from Blastochloris viridis
PLOS ONE, 2012Co-Authors: Weixiao Yuan Wahlgren, Hadil Omran, David Von Stetten, Antoine Royant, Sjoerd Van Der Post, Gergely KatonaAbstract:Iron storage and elimination of toxic ferrous iron are the responsibility of bacterioferritins in bacterial species. Bacterioferritins are capable of oxidizing iron using molecular oxygen and import iron ions into the large central cavity of the protein, where they are stored in a mineralized form. We isolated, crystallized bacterioferritin from the microaerophilic/anaerobic, purple non-sulfur bacterium Blastochloris viridis and determined its amino acid sequence and X-ray structure. The structure and sequence revealed similarity to other purple bacterial species with substantial differences in the pore regions. Static 3- and 4-fold pores do not allow the passage of iron ions even though structural dynamics may assist the iron gating. On the other hand the B-pore is open to water and larger ions in its native state. In order to study the mechanism of iron import, multiple soaking experiments were performed. Upon Fe(II) and urea treatment the ferroxidase site undergoes reorganization as seen in bacterioferritin from Escherichia coli and Pseudomonas aeruginosa. When soaking with Fe(II) only, a closely bound small molecular ligand is observed close to Fe1 and the coordination of Glu94 to Fe2 changes from bidentate to monodentate. DFT calculations indicate that the bound ligand is most likely a water or a hydroxide molecule representing a product complex. On the other hand the different soaking treatments did not modify the conformation of other pore regions.
Linda C. Johansson - One of the best experts on this subject based on the ideXlab platform.
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From Macrocrystals to Microcrystals: A Strategy for Membrane Protein Serial Crystallography
Structure (London England : 1993), 2017Co-Authors: Robert Dods, Petra Båth, David Arnlund, Erik Malmerberg, Kenneth R. Beyerlein, Garrett Nelson, Mengling Liang, Rajiv Harimoorthy, Peter Berntsen, Linda C. JohanssonAbstract:Summary Serial protein crystallography was developed at X-ray free-electron lasers (XFELs) and is now also being applied at storage ring facilities. Robust strategies for the growth and optimization of microcrystals are needed to advance the field. Here we illustrate a generic strategy for recovering high-density homogeneous samples of microcrystals starting from conditions known to yield large (macro) crystals of the photosynthetic reaction center of Blastochloris viridis (RC vir ). We first crushed these crystals prior to multiple rounds of microseeding. Each cycle of microseeding facilitated improvements in the RC vir serial femtosecond crystallography (SFX) structure from 3.3-A to 2.4-A resolution. This approach may allow known crystallization conditions for other proteins to be adapted to exploit novel scientific opportunities created by serial crystallography.
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Asymmetry in serial femtosecond crystallography data
Acta Crystallographica Section A Foundations and Advances, 2017Co-Authors: Amit Sharma, Weixiao Yuan Wahlgren, Linda C. Johansson, Richard Neutze, Elin Dunevall, Gergely KatonaAbstract:Serial crystallography is an increasingly important approach to protein crystallography that exploits both X-ray free-electron laser (XFEL) and synchrotron radiation. Serial crystallography recovers complete X-ray diffraction data by processing and merging diffraction images from thousands of randomly oriented non-uniform microcrystals, of which all observations are partial Bragg reflections. Random fluctuations in the XFEL pulse energy spectrum, variations in the size and shape of microcrystals, integrating over millions of weak partial observations and instabilities in the XFEL beam position lead to new types of experimental errors. The quality of Bragg intensity estimates deriving from serial crystallography is therefore contingent upon assumptions made while modeling these data. Here it is observed that serial femtosecond crystallography (SFX) Bragg reflections do not follow a unimodal Gaussian distribution and it is recommended that an idealized assumption of single Gaussian peak profiles be relaxed to incorporate apparent asymmetries when processing SFX data. The phenomenon is illustrated by re-analyzing data collected from microcrystals of the Blastochloris viridis photosynthetic reaction center and comparing these intensity observations with conventional synchrotron data. The results show that skewness in the SFX observations captures the essence of the Wilson plot and an empirical treatment is suggested that can help to separate the diffraction Bragg intensity from the background.
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography
2016Co-Authors: Linda C. Johansson, Gergely Katona, David Arnlund, Thomas A. White, Daniel P. Deponte, Robert L. Shoeman, Anton Barty, Cecilia Wickstr, Amit Sharma, Garth J. WilliamsAbstract:Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8Å resolution and determine its serial femtosecond crystallography structure to 3.5Å resolution. Although every microcrystal is exposed to a dose of 33MGy, no signs of X-ray-induced radiation damage are visible in this integra
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Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser
Nature Methods, 2014Co-Authors: David Arnlund, Cecilia Wickstrand, Linda C. Johansson, Daniel P. Deponte, Erik Malmerberg, Anton Barty, Garth J. Williams, Jan Davidsson, Despina Milathianaki, Robert L. ShoemanAbstract:We describe a method to measure ultrafast protein structural changes using time-resolved wide-angle X-ray scattering at an X-ray free-electron laser. We demonstrated this approach using multiphoton excitation of the Blastochloris viridis photosynthetic reaction center, observing an ultrafast global conformational change that arises within picoseconds and precedes the propagation of heat through the protein. This provides direct structural evidence for a 'protein quake': the hypothesis that proteins rapidly dissipate energy through quake-like structural motions. A 'protein quake' is directly monitored on the picosecond timescale using the method of time-resolved wide-angle X-ray scattering at an X-ray free-electron laser.
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Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser.
Nature methods, 2014Co-Authors: David Arnlund, Cecilia Wickstrand, Linda C. Johansson, Daniel P. Deponte, Erik Malmerberg, Anton Barty, Garth J. Williams, Jan Davidsson, Despina Milathianaki, Robert L. ShoemanAbstract:We describe a method to measure ultrafast protein structural changes using time-resolved wide-angle X-ray scattering at an X-ray free-electron laser. We demonstrated this approach using multiphoton excitation of the Blastochloris viridis photosynthetic reaction center, observing an ultrafast global conformational change that arises within picoseconds and precedes the propagation of heat through the protein. This provides direct structural evidence for a 'protein quake': the hypothesis that proteins rapidly dissipate energy through quake-like structural motions.
David Arnlund - One of the best experts on this subject based on the ideXlab platform.
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Ultrafast structural changes within a photosynthetic reaction centre
Nature, 2021Co-Authors: Robert Dods, Petra Båth, Dmitry Morozov, Viktor Ahlberg Gagnér, David Arnlund, Hoi Ling Luk, Joachim Kübel, Michał Maj, Adams Vallejos, Cecilia WickstrandAbstract:Time-resolved serial femtosecond crystallography is used to reveal the structural changes that stabilize the charge-separation steps of electron-transfer reactions in the photosynthetic reaction centre of Blastochloris viridis on a timescale of picoseconds. Photosynthetic reaction centres harvest the energy content of sunlight by transporting electrons across an energy-transducing biological membrane. Here we use time-resolved serial femtosecond crystallography^ 1 using an X-ray free-electron laser^ 2 to observe light-induced structural changes in the photosynthetic reaction centre of Blastochloris viridis on a timescale of picoseconds. Structural perturbations first occur at the special pair of chlorophyll molecules of the photosynthetic reaction centre that are photo-oxidized by light. Electron transfer to the menaquinone acceptor on the opposite side of the membrane induces a movement of this cofactor together with lower amplitude protein rearrangements. These observations reveal how proteins use conformational dynamics to stabilize the charge-separation steps of electron-transfer reactions.
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Ultrafast structural changes within a photosynthetic reaction centre.
Nature, 2020Co-Authors: Robert Dods, Petra Båth, Dmitry Morozov, Viktor Ahlberg Gagnér, David Arnlund, Hoi Ling Luk, Joachim Kübel, Michał Maj, Adams Vallejos, Cecilia WickstrandAbstract:Photosynthetic reaction centres harvest the energy content of sunlight by transporting electrons across an energy-transducing biological membrane. Here we use time-resolved serial femtosecond crystallography1 using an X-ray free-electron laser2 to observe light-induced structural changes in the photosynthetic reaction centre of Blastochloris viridis on a timescale of picoseconds. Structural perturbations first occur at the special pair of chlorophyll molecules of the photosynthetic reaction centre that are photo-oxidized by light. Electron transfer to the menaquinone acceptor on the opposite side of the membrane induces a movement of this cofactor together with lower amplitude protein rearrangements. These observations reveal how proteins use conformational dynamics to stabilize the charge-separation steps of electron-transfer reactions.
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From Macrocrystals to Microcrystals: A Strategy for Membrane Protein Serial Crystallography
Structure (London England : 1993), 2017Co-Authors: Robert Dods, Petra Båth, David Arnlund, Erik Malmerberg, Kenneth R. Beyerlein, Garrett Nelson, Mengling Liang, Rajiv Harimoorthy, Peter Berntsen, Linda C. JohanssonAbstract:Summary Serial protein crystallography was developed at X-ray free-electron lasers (XFELs) and is now also being applied at storage ring facilities. Robust strategies for the growth and optimization of microcrystals are needed to advance the field. Here we illustrate a generic strategy for recovering high-density homogeneous samples of microcrystals starting from conditions known to yield large (macro) crystals of the photosynthetic reaction center of Blastochloris viridis (RC vir ). We first crushed these crystals prior to multiple rounds of microseeding. Each cycle of microseeding facilitated improvements in the RC vir serial femtosecond crystallography (SFX) structure from 3.3-A to 2.4-A resolution. This approach may allow known crystallization conditions for other proteins to be adapted to exploit novel scientific opportunities created by serial crystallography.
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography
2016Co-Authors: Linda C. Johansson, Gergely Katona, David Arnlund, Thomas A. White, Daniel P. Deponte, Robert L. Shoeman, Anton Barty, Cecilia Wickstr, Amit Sharma, Garth J. WilliamsAbstract:Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8Å resolution and determine its serial femtosecond crystallography structure to 3.5Å resolution. Although every microcrystal is exposed to a dose of 33MGy, no signs of X-ray-induced radiation damage are visible in this integra
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X-ray free-electron laser based methods for structural and ultrafast dynamics studies of a photosynthetic reaction centre
2014Co-Authors: David ArnlundAbstract:Life on earth is fuelled by the energy of sunlight, which must first be captured and converted into a chemical energy form useful to the cell. This process is known as photosynthesis and the major pathway of this energy conversion is via photosynthetic reaction centres. These enzymes convert the energy content of an absorbed photon into a transmembrane potential difference via the movements of electrons. Increasing our knowledge of the three-dimensional fold and structural changes that takes place within photosynthetic reaction centres is therefore of considerable importance for understanding biological photosynthesis. The aim of this work has been to adapt methods for both crystallographic and solution phase structural studies of membrane proteins to the unique properties of X-ray freeelectron laser (XFEL) radiation. To accomplish this, a new crystallization technique for the photosynthetic reaction centre from the purple bacterium Blastochloris viridis (RCvir) was developed which was suitable for serial femtosecond crystallography (SFX) experiments at an XFEL. Our initial experiments at the Linac Coherent Light Source (LCLS), the world’s first XFEL, yielded an SFX structure of RCvir to 8.2 A resolution. After the LCLS decreased the X-ray wavelength at which the facility could operate, and in combination with improved crystallization conditions, we later resolved the SFX structure of RCvir to 3.5 A resolution. Whether or not ultrafast structural changes in RCvir occur in photosynthesis has been debated for two decades. We addressed this question by developing time-resolved wide-angle X-ray scattering (TR-WAXS) studies at the LCLS that could capture rapid structural changes in solubilized samples of RCvir. Proof-of-principle experiments revealed a structural deformation that propagated through the RCvir protein following multi-photon absorption by its cofactors, enabling a protein quake through a photosynthetic protein to be visualized. Further insight was provided by a second TRWAXS experiment in which this structural signal was observed in the data as the pump laser fluence was decreased to less than one photon absorbed per RCvir molecule. This result implies that, even under physiological conditions of normal sunlight, ultrafast protein structural rearrangements may influence the primary charge separation events of biological photosynthesis.
Hitoshi Tamiaki - One of the best experts on this subject based on the ideXlab platform.
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Complete Genome Sequence of the Bacteriochlorophyll b-Producing Photosynthetic Bacterium Blastochloris viridis.
Genome announcements, 2015Co-Authors: Yusuke Tsukatani, Yuu Hirose, Jiro Harada, Naomi Misawa, Keita Mori, Kazuhito Inoue, Hitoshi TamiakiAbstract:We report the complete genome sequence of the purple photosynthetic bacterium Blastochloris viridis belonging to α-Proteobacteria. This is the first completed genome sequence of a phototroph producing bacteriochlorophyll b. The genome information will be useful for further analysis of the photosynthetic energy conversion system and bacteriochlorophyll pigment biosynthesis.
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Rhodobacter sphaeroides mutants overexpressing chlorophyllide a oxidoreductase of Blastochloris viridis elucidate functions of enzymes in late bacteriochlorophyll biosynthetic pathways
Scientific reports, 2015Co-Authors: Yusuke Tsukatani, Jiro Harada, Jiro Nomata, Haruki Yamamoto, Yuichi Fujita, Tadashi Mizoguchi, Hitoshi TamiakiAbstract:Rhodobacter sphaeroides mutants overexpressing chlorophyllide a oxidoreductase of Blastochloris viridis elucidate functions of enzymes in late bacteriochlorophyll biosynthetic pathways
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Rhodobacter sphaeroides mutants overexpressing chlorophyllide a oxidoreductase of Blastochloris viridis elucidate functions of enzymes in late bacteriochlorophyll biosynthetic pathways
Scientific Reports, 2015Co-Authors: Yusuke Tsukatani, Jiro Harada, Jiro Nomata, Haruki Yamamoto, Yuichi Fujita, Tadashi Mizoguchi, Hitoshi TamiakiAbstract:In previous studies we have demonstrated that chlorophyllide a oxidoreductases (CORs) from bacteriochlorophyll (BChl) a -producing Rhodobacter species and BChl b -producing Blastochloris viridis show distinct substrate recognition and different catalytic hydrogenation reactions and that these two types of CORs therefore cause committed steps for BChls a and b biosynthesis. In this study, COR genes from B. viridis were incorporated and overexpressed in a series of Rhodobacter sphaeroides mutants. We found that the following two factors are essential in making R. sphaeroides produce BChl b : the loss of functions of both intrinsic COR and 8-vinyl reductase (BciA) in the host R. sphaeroides strain; and expression of the BchYZ catalytic components of COR from B. viridis , not the complete set of COR (BchXYZ), in the host strain. In addition, we incorporated bchYZ of B. viridis into the R. sphaeroides mutant lacking BchJ and BciA, resulting in the strain accumulating both BChl a and BChl b . This is the first example of an anoxygenic photosynthetic bacterium producing BChls a and b together. The results suggest that BchJ enhances activity of the intrinsic COR. The physiological significance of BchJ in pigment biosynthetic pathways will be discussed.
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An unexpectedly branched biosynthetic pathway for bacteriochlorophyll b capable of absorbing near-infrared light
Scientific Reports, 2013Co-Authors: Yusuke Tsukatani, Jiro Harada, Jiro Nomata, Haruki Yamamoto, Yuichi Fujita, Tadashi Mizoguchi, Taichi Yoshitomi, Masahiro Kasahara, Hitoshi TamiakiAbstract:Chlorophyllous pigments are essential for photosynthesis. Bacteriochlorophyll (BChl) b has the characteristic C8-ethylidene group and therefore is the sole naturally occurring pigment having an absorption maximum at near-infrared light wavelength. Here we report that chlorophyllide a oxidoreductase (COR), a nitrogenase-like enzyme, showed distinct substrate recognition and catalytic reaction between BChl a- and b -producing proteobacteria. COR from BChl b -producing Blastochloris viridis synthesized the C8-ethylidene group from 8-vinyl-chlorophyllide a . In contrast, despite the highly conserved primary structures, COR from BChl a -producing Rhodobacter capsulatus catalyzes the C8-vinyl reduction as well as the previously known reaction of the C7 = C8 double bond reduction on 8-vinyl-chlorophyllide a . The present data indicate that the plasticity of the nitrogenase-like enzyme caused the branched pathways of BChls a and b biosynthesis, ultimately leading to ecologically different niches of BChl a - and b -based photosynthesis differentiated by more than 150 nm wavelength.
James R. Norris - One of the best experts on this subject based on the ideXlab platform.
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Electronic structure of the primary electron donor of Blastochloris viridis heterodimer mutants : high field EPR study.
Biochimica et biophysica acta, 2010Co-Authors: Nina Ponomarenko, Oleg G. Poluektov, E.j. Bylina, James R. NorrisAbstract:Abstract High-field electron paramagnetic resonance (HF EPR) has been employed to investigate the primary electron donor electronic structure of Blastochloris viridis heterodimer mutant reaction centers (RCs). In these mutants the amino acid substitution His(M200)Leu or His(L173)Leu eliminates a ligand to the primary electron donor, resulting in the loss of a magnesium in one of the constituent bacteriochlorophylls (BChl). Thus, the native BChl/BChl homodimer primary donor is converted into a BChl/bacteriopheophytin (BPhe) heterodimer. The heterodimer primary donor radical in chemically oxidized RCs exhibits a broadened EPR line indicating a highly asymmetric distribution of the unpaired electron over both dimer constituents. Observed triplet state EPR signals confirm localization of the excitation on the BChl half of the heterodimer primary donor. Theoretical simulation of the triplet EPR lineshapes clearly shows that, in the case of mutants, triplet states are formed by an intersystem crossing mechanism in contrast to the radical pair mechanism in wild type RCs. Photooxidation of the mutant RCs results in formation of a BPhe anion radical within the heterodimer pair. The accumulation of an intradimer BPhe anion is caused by the substantial loss of interaction between constituents of the heterodimer primary donor along with an increase in the reduction potential of the heterodimer primary donor D/D+ couple. This allows oxidation of the cytochrome even at cryogenic temperatures and reduction of each constituent of the heterodimer primary donor individually. Despite a low yield of primary donor radicals, the enhancement of the semiquinone–iron pair EPR signals in these mutants indicates the presence of kinetically viable electron donors.
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Photoactivation of the Photosynthetic Reaction Center of Blastochloris viridis in the Crystalline State
The journal of physical chemistry. B, 2006Co-Authors: Richard H. G. Baxter, Elmars Krausz, James R. NorrisAbstract:Photoactivation in crystals of the bacterial reaction center of Blastochloris viridis was investigated by near-infrared spectroscopy. The bleaching of the special pair absorption at 970 nm and the simultaneous rise of the special pair cation absorption at 1300 nm were measured in response to transient irradiation by a HeNe laser over 5 orders of magnitude in laser power. The resulting power-saturation curve can be used to estimate the true extent of photoactivation achieved in a prior time-resolved crystallographic experiment (Baxter et al. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 5982−5987). The overall extent of photoactivation was 50%, which demonstrates that the time-resolved crystallographic method can be applied to the optically dense reaction center crystals. Measurement of the charge-recombination rate, however, suggests the presence of a long-lived P+ state within the crystal.
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Cryogenic structure of the photosynthetic reaction center of Blastochloris viridis in the light and dark.
Acta Crystallographica Section D Biological Crystallography, 2005Co-Authors: Richard H. G. Baxter, Brandon Luke Seagle, Nina Ponomarenko, James R. NorrisAbstract:The structure of the Blastochloris viridis photosynthetic reaction center has been determined at 100 K by flash-freezing crystals. A data set to 2.2 A resolution provides a well determined model of the wild-type protein. Of particular interest are the position, occupancy and heterogeneity of the Q(B)-binding site. Data were also collected from a crystal frozen immediately after illumination. The data support predominant binding of Q(B) in the proximal position in both the neutral and charge-separated states.
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Specific radiation damage illustrates light-induced structural changes in the photosynthetic reaction center.
Journal of the American Chemical Society, 2004Co-Authors: Richard H. G. Baxter, Nina Ponomarenko, Brandon-luke L. Seagle, James R. NorrisAbstract:The photosynthetic reaction center of the purple non-sulfur bacterium Blastochloris viridis was frozen in the presence and absence of illumination. Differences in the resulting datasets are monitored using the difference Fourier method. Radiation damage is localized to those parts of the protein that are significant for electron transfer, and show changes that are sensitive to oxidation and protonation state.
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Time-resolved crystallographic studies of light-induced structural changes in the photosynthetic reaction center
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Richard H. G. Baxter, Nina Ponomarenko, Vukica Šrajer, R. Pahl, Keith Moffat, James R. NorrisAbstract:Light-induced structural changes in the bacterial reaction center were studied by a time-resolved crystallographic experiment. Crystals of protein from Blastochloris viridis (formerly Rhodopseudomonas viridis) were reconstituted with ubiquinone and analyzed by monochromatic and Laue diffraction, in the dark and 3 ms after illuminating the crystal with a pulsed laser (630 nm, 3 mJ/pulse, 7 ns duration). Refinement of monochromatic data shows that ubiquinone binds only in the “proximal” QB binding site. No significant structural difference was observed between the light and dark datasets; in particular, no quinone motion was detected. This result may be reconciled with previous studies by postulating equilibration of the “distal” and “proximal” binding sites upon extended dark adaption, and in which movement of ubiquinone is not the conformational gate for the first electron transfer between QA and QB.