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Taking snapshots of photosynthetic water oxidation using femtosecond X-ray diffraction and spectroscopy.

Kern J, Tran R, Alonso-Mori R, Koroidov S, Echols N, Hattne J, Ibrahim M, Gul S, Laksmono H, Sierra RG, Gildea RJ, Han G, Hellmich J, Lassalle-Kaiser B, Chatterjee R, Brewster AS, Stan CA, Glöckner C, Lampe A, DiFiore D, Milathianaki D, Fry AR, Seibert MM, Koglin JE, Gallo E, Uhlig J, Sokaras D, Weng TC, Zwart PH, Skinner DE, Bogan MJ, Messerschmidt M, Glatzel P, Williams GJ, Boutet S, Adams PD, Zouni A, Messinger J, Sauter NK, Bergmann U, Yano J, Yachandra VK - Nat Commun (2014)

Bottom Line: The spectra show that the initial O-O bond formation, coupled to Mn reduction, does not yet occur within 250 μs after the third flash.Diffraction data of all states studied exhibit an anomalous scattering signal from Mn but show no significant structural changes at the present resolution of 4.5 Å.This study represents the initial frames in a molecular movie of the structural changes during the catalytic reaction in photosystem II.

View Article: PubMed Central - PubMed

Affiliation: 1] Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA [2] LCLS, SLAC National Accelerator Laboratory, Menlo Park, California 94025, USA.

ABSTRACT
The dioxygen we breathe is formed by light-induced oxidation of water in photosystem II. O2 formation takes place at a catalytic manganese cluster within milliseconds after the photosystem II reaction centre is excited by three single-turnover flashes. Here we present combined X-ray emission spectra and diffraction data of 2-flash (2F) and 3-flash (3F) photosystem II samples, and of a transient 3F' state (250 μs after the third flash), collected under functional conditions using an X-ray free electron laser. The spectra show that the initial O-O bond formation, coupled to Mn reduction, does not yet occur within 250 μs after the third flash. Diffraction data of all states studied exhibit an anomalous scattering signal from Mn but show no significant structural changes at the present resolution of 4.5 Å. This study represents the initial frames in a molecular movie of the structural changes during the catalytic reaction in photosystem II.

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Anomalous signal from Mn for different illumination states of PS II A) Anomalous map of the OEC in PS II is shown for the 2F data (magenta) in monomer I. (B) Anomalous map of the 3F data in monomer I. (C) Anomalous map of the 2F (cyan) and 3F (magenta) data in monomer I, orientation is rotated by 90° around horizontal and vertical axis compared to the view in A. (D) Anomalous map for monomer II, 2F (cyan) and 3F (magenta) data are shown, view direction is similar to panel C. All maps shown are anomalous difference simulated annealing omit maps contoured at 3σ.
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Figure 6: Anomalous signal from Mn for different illumination states of PS II A) Anomalous map of the OEC in PS II is shown for the 2F data (magenta) in monomer I. (B) Anomalous map of the 3F data in monomer I. (C) Anomalous map of the 2F (cyan) and 3F (magenta) data in monomer I, orientation is rotated by 90° around horizontal and vertical axis compared to the view in A. (D) Anomalous map for monomer II, 2F (cyan) and 3F (magenta) data are shown, view direction is similar to panel C. All maps shown are anomalous difference simulated annealing omit maps contoured at 3σ.

Mentions: In PS II, a clear anomalous signal (Fig. 5C, D, Fig. 6 and Supplementary Fig. 8 and 9) from Mn in the OEC is also detected in all four data sets (0F, 2F, 3F, and 3F′) (Table S1). Figs. 5C,D show the anomalous difference map from the 3F data after omitting the OEC and performing simulated annealing refinement. It is evident from the overview shown in Fig. 5C that the largest peak (σ > 6) in the anomalous density is located at the position of the OEC. The density covers the Mn ions in the cluster and does not include the Ca (Fig. 5D) as expected from the weaker anomalous contribution of Ca at 7.1 keV (f″ of 1.6 for Ca compared to 3.4 for Mn at 7.1 keV). Similar results were obtained for the other PS II datasets for both monomers in the PS II dimer (Fig. 6 and Supplementary Fig. 8 and 9). It should ne noted however that the anomalous difference Patterson maps did not reveal peaks above the noise level attributable to Mn. This result is expected as also the anomalous data measured at SR sources at 3.5 Å resolution10 did not yield any peaks in the Patterson map above the noise level, due to the large protein mass and the low number of anomalous scatterers per unit cell volume.


Taking snapshots of photosynthetic water oxidation using femtosecond X-ray diffraction and spectroscopy.

Kern J, Tran R, Alonso-Mori R, Koroidov S, Echols N, Hattne J, Ibrahim M, Gul S, Laksmono H, Sierra RG, Gildea RJ, Han G, Hellmich J, Lassalle-Kaiser B, Chatterjee R, Brewster AS, Stan CA, Glöckner C, Lampe A, DiFiore D, Milathianaki D, Fry AR, Seibert MM, Koglin JE, Gallo E, Uhlig J, Sokaras D, Weng TC, Zwart PH, Skinner DE, Bogan MJ, Messerschmidt M, Glatzel P, Williams GJ, Boutet S, Adams PD, Zouni A, Messinger J, Sauter NK, Bergmann U, Yano J, Yachandra VK - Nat Commun (2014)

Anomalous signal from Mn for different illumination states of PS II A) Anomalous map of the OEC in PS II is shown for the 2F data (magenta) in monomer I. (B) Anomalous map of the 3F data in monomer I. (C) Anomalous map of the 2F (cyan) and 3F (magenta) data in monomer I, orientation is rotated by 90° around horizontal and vertical axis compared to the view in A. (D) Anomalous map for monomer II, 2F (cyan) and 3F (magenta) data are shown, view direction is similar to panel C. All maps shown are anomalous difference simulated annealing omit maps contoured at 3σ.
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Related In: Results  -  Collection

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Show All Figures
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Figure 6: Anomalous signal from Mn for different illumination states of PS II A) Anomalous map of the OEC in PS II is shown for the 2F data (magenta) in monomer I. (B) Anomalous map of the 3F data in monomer I. (C) Anomalous map of the 2F (cyan) and 3F (magenta) data in monomer I, orientation is rotated by 90° around horizontal and vertical axis compared to the view in A. (D) Anomalous map for monomer II, 2F (cyan) and 3F (magenta) data are shown, view direction is similar to panel C. All maps shown are anomalous difference simulated annealing omit maps contoured at 3σ.
Mentions: In PS II, a clear anomalous signal (Fig. 5C, D, Fig. 6 and Supplementary Fig. 8 and 9) from Mn in the OEC is also detected in all four data sets (0F, 2F, 3F, and 3F′) (Table S1). Figs. 5C,D show the anomalous difference map from the 3F data after omitting the OEC and performing simulated annealing refinement. It is evident from the overview shown in Fig. 5C that the largest peak (σ > 6) in the anomalous density is located at the position of the OEC. The density covers the Mn ions in the cluster and does not include the Ca (Fig. 5D) as expected from the weaker anomalous contribution of Ca at 7.1 keV (f″ of 1.6 for Ca compared to 3.4 for Mn at 7.1 keV). Similar results were obtained for the other PS II datasets for both monomers in the PS II dimer (Fig. 6 and Supplementary Fig. 8 and 9). It should ne noted however that the anomalous difference Patterson maps did not reveal peaks above the noise level attributable to Mn. This result is expected as also the anomalous data measured at SR sources at 3.5 Å resolution10 did not yield any peaks in the Patterson map above the noise level, due to the large protein mass and the low number of anomalous scatterers per unit cell volume.

Bottom Line: The spectra show that the initial O-O bond formation, coupled to Mn reduction, does not yet occur within 250 μs after the third flash.Diffraction data of all states studied exhibit an anomalous scattering signal from Mn but show no significant structural changes at the present resolution of 4.5 Å.This study represents the initial frames in a molecular movie of the structural changes during the catalytic reaction in photosystem II.

View Article: PubMed Central - PubMed

Affiliation: 1] Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA [2] LCLS, SLAC National Accelerator Laboratory, Menlo Park, California 94025, USA.

ABSTRACT
The dioxygen we breathe is formed by light-induced oxidation of water in photosystem II. O2 formation takes place at a catalytic manganese cluster within milliseconds after the photosystem II reaction centre is excited by three single-turnover flashes. Here we present combined X-ray emission spectra and diffraction data of 2-flash (2F) and 3-flash (3F) photosystem II samples, and of a transient 3F' state (250 μs after the third flash), collected under functional conditions using an X-ray free electron laser. The spectra show that the initial O-O bond formation, coupled to Mn reduction, does not yet occur within 250 μs after the third flash. Diffraction data of all states studied exhibit an anomalous scattering signal from Mn but show no significant structural changes at the present resolution of 4.5 Å. This study represents the initial frames in a molecular movie of the structural changes during the catalytic reaction in photosystem II.

Show MeSH
Related in: MedlinePlus