| 1. |
- Alonso-Mori, Roberto, et al.
(författare)
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Energy-dispersive X-ray emission spectroscopy using an X-ray free-electron laser in a shot-by-shot mode
- 2012
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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. ; 109:47, s. 19103-19107
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Tidskriftsartikel (refereegranskat)abstract
- The ultrabright femtosecond X-ray pulses provided by X-ray free-electron lasers open capabilities for studying the structure and dynamics of a wide variety of systems beyond what is possible with synchrotron sources. Recently, this probe-before-destroy approach has been demonstrated for atomic structure determination by serial X-ray diffraction of microcrystals. There has been the question whether a similar approach can be extended to probe the local electronic structure by X-ray spectroscopy. To address this, we have carried out femtosecond X-ray emission spectroscopy (XES) at the Linac Coherent Light Source using redox-active Mn complexes. XES probes the charge and spin states as well as the ligand environment, critical for understanding the functional role of redox-active metal sites. K beta(1,3) XES spectra of Mn-II and Mn-2(III,IV) complexes at room temperature were collected using a wavelength dispersive spectrometer and femtosecond X-ray pulses with an individual dose of up to > 100 MGy. The spectra were found in agreement with undamaged spectra collected at low dose using synchrotron radiation. Our results demonstrate that the intact electronic structure of redox active transition metal compounds in different oxidation states can be characterized with this shot-by-shot method. This opens the door for studying the chemical dynamics of metal catalytic sites by following reactions under functional conditions. The technique can be combined with X-ray diffraction to simultaneously obtain the geometric structure of the overall protein and the local chemistry of active metal sites and is expected to prove valuable for understanding the mechanism of important metalloproteins, such as photosystem II.
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| 2. |
- Esmaeildoost, Niloofar, et al.
(författare)
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Heterogeneous Ice Growth in Micron-Sized Water Droplets Due to Spontaneous Freezing
- 2022
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Ingår i: Crystals. - : MDPI AG. - 2073-4352. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Understanding how ice nucleates and grows into larger crystals is of crucial importance for many research fields. The purpose of this study was to shed light on the phase and structure of ice once a nucleus is formed inside a metastable water droplet. Wide-angle X-ray scattering (WAXS) was performed on micron-sized droplets evaporatively cooled to temperatures where homogeneous nucleation occurs. We found that for our weak hits ice grows more cubic compared to the strong hits that are completely hexagonal. Due to efficient heat removal caused by evaporation, we propose that the cubicity of ice at the vicinity of the droplet’s surface is higher than for ice formed within the bulk of the droplet. Moreover, the Bragg peaks were classified based on their geometrical shapes and positions in reciprocal space, which showed that ice grows heterogeneously with a significant population of peaks indicative of truncation rods and crystal defects. Frequent occurrences of the (100) reflection with extended in-planar structure suggested that large planar ice crystals form at the droplet surface, then fracture into smaller domains to accommodate to the curvature of the droplets. Planar faulting due to misaligned domains would explain the increased cubicity close to the droplet surface.
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| 3. |
- Hattne, Johan, et al.
(författare)
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Accurate macromolecular structures using minimal measurements from X-ray free-electron lasers
- 2014
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Ingår i: Nature Methods. - 1548-7091 .- 1548-7105. ; 11:5, s. 545-548
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Tidskriftsartikel (refereegranskat)abstract
- X-ray free-electron laser (XFEL) sources enable the use of crystallography to solve three-dimensional macromolecular structures under native conditions and without radiation damage. Results to date, however, have been limited by the challenge of deriving accurate Bragg intensities from a heterogeneous population of microcrystals, while at the same time modeling the X-ray spectrum and detector geometry. Here we present a computational approach designed to extract meaningful high-resolution signals from fewer diffraction measurements.
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| 4. |
- Hattne, Johan, et al.
(författare)
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Accurate macromolecular structures using minimal measurements from X-ray free-electron lasers
- 2014
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Ingår i: Nature Methods. - : Springer Science and Business Media LLC. - 1548-7091 .- 1548-7105. ; 11:5, s. 545-548
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Tidskriftsartikel (refereegranskat)abstract
- X-ray free-electron laser (XFEL) sources enable the use of crystallography to solve three-dimensional macromolecular structures under native conditions and without radiation damage. Results to date, however, have been limited by the challenge of deriving accurate Bragg intensities from a heterogeneous population of microcrystals, while at the same time modeling the X-ray spectrum and detector geometry. Here we present a computational approach designed to extract meaningful high-resolution signals from fewer diffraction measurements.
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| 5. |
- Kern, Jan, et al.
(författare)
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Room temperature femtosecond X-ray diffraction of photosystem II microcrystals
- 2012
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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. ; 109:25, s. 9721-9726
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Tidskriftsartikel (refereegranskat)abstract
- Most of the dioxygen on earth is generated by the oxidation of water by photosystem II (PS II) using light from the sun. This lightdriven, four-photon reaction is catalyzed by the Mn4CaO5 cluster located at the lumenal side of PS II. Various X-ray studies have been carried out at cryogenic temperatures to understand the intermediate steps involved in the water oxidation mechanism. However, the necessity for collecting data at room temperature, especially for studying the transient steps during the O-O bond formation, requires the development of new methodologies. In this paper we report room temperature X-ray diffraction data of PS II microcrystals obtained using ultrashort (<50 fs) 9 keV X-ray pulses from a hard X-ray free electron laser, namely the Linac Coherent Light Source. The results presented here demonstrate that the probe before destroy approach using an X-ray free electron laser works even for the highly-sensitive Mn4CaO5 cluster in PS II at room temperature. We show that these data are comparable to those obtained in synchrotron radiation studies as seen by the similarities in the overall structure of the helices, the protein subunits and the location of the various cofactors. This work is, therefore, an important step toward future studies for resolving the structure of the Mn4CaO5 cluster without any damage at room temperature, and of the reaction intermediates of PS II during O-O bond formation.
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| 6. |
- Kern, Jan, et al.
(författare)
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Simultaneous Femtosecond X-ray Spectroscopy and Diffraction of Photosystem II at Room Temperature
- 2013
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 340:6131, s. 491-495
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Tidskriftsartikel (refereegranskat)abstract
- Intense femtosecond x-ray pulses produced at the Linac Coherent Light Source (LCLS) were used for simultaneous x-ray diffraction (XRD) and x-ray emission spectroscopy (XES) of microcrystals of photosystem II (PS II) at room temperature. This method probes the overall protein structure and the electronic structure of the Mn4CaO5 cluster in the oxygen-evolving complex of PS II. XRD data are presented from both the dark state (S-1) and the first illuminated state (S-2) of PS II. Our simultaneous XRD-XES study shows that the PS II crystals are intact during our measurements at the LCLS, not only with respect to the structure of PS II, but also with regard to the electronic structure of the highly radiation-sensitive Mn4CaO5 cluster, opening new directions for future dynamics studies.
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| 7. |
- Kern, Jan, et al.
(författare)
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Taking snapshots of photosynthetic water oxidation using femtosecond X-ray diffraction and spectroscopy
- 2014
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 5, s. 4371-
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Tidskriftsartikel (refereegranskat)abstract
- The dioxygen we breathe is formed by light-induced oxidation of water in photosystem II. O-2 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 mu s 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 mu s 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 angstrom. 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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| 8. |
- Kubin, Markus, et al.
(författare)
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Soft x-ray absorption spectroscopy of metalloproteins and high-valent metal-complexes at room temperature using free-electron lasers
- 2017
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Ingår i: Structural Dynamics. - : AMER INST PHYSICS. - 2329-7778. ; 4:5
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Tidskriftsartikel (refereegranskat)abstract
- X-ray absorption spectroscopy at the L-edge of 3d transition metals provides unique information on the local metal charge and spin states by directly probing 3d-derived molecular orbitals through 2p-3d transitions. However, this soft x-ray technique has been rarely used at synchrotron facilities for mechanistic studies of metalloenzymes due to the difficulties of x-ray-induced sample damage and strong background signals from light elements that can dominate the low metal signal. Here, we combine femtosecond soft x-ray pulses from a free-electron laser with a novel x-ray fluorescence-yield spectrometer to overcome these difficulties. We present L-edge absorption spectra of inorganic high-valent Mn complexes (Mn similar to 6-15 mmol/l) with no visible effects of radiation damage. We also present the first L-edge absorption spectra of the oxygen evolving complex (Mn4CaO5) in Photosystem II (Mn < 1 mmol/l) at room temperature, measured under similar conditions. Our approach opens new ways to study metalloenzymes under functional conditions. (C) 2017 Author(s).
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| 9. |
- Kupitz, Christopher, et al.
(författare)
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Serial time-resolved crystallography of photosystem II using a femtosecond X-ray laser
- 2014
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 513:7517, s. 261-265
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Tidskriftsartikel (refereegranskat)abstract
- Photosynthesis, a process catalysed by plants, algae and cyanobacteria converts sunlight to energy thus sustaining all higher life on Earth. Two large membrane protein complexes, photosystem I and II (PSI and PSII), act in series to catalyse the light-driven reactions in photosynthesis. PSII catalyses the light-driven water splitting process, which maintains the Earth's oxygenic atmosphere. In this process, the oxygen-evolving complex (OEC) of PSII cycles through five states, S0 to S4, in which four electrons are sequentially extracted from the OEC in four light-driven charge-separation events. Here we describe time resolved experiments on PSII nano/microcrystals from Thermosynechococcus elongatus performed with the recently developed technique of serial femtosecond crystallography. Structures have been determined from PSII in the dark S1 state and after double laser excitation (putative S3 state) at 5 and 5.5 Å resolution, respectively. The results provide evidence that PSII undergoes significant conformational changes at the electron acceptor side and at the Mn4CaO5 core of the OEC. These include an elongation of the metal cluster, accompanied by changes in the protein environment, which could allow for binding of the second substrate water molecule between the more distant protruding Mn (referred to as the 'dangler' Mn) and the Mn3CaOx cubane in the S2 to S3 transition, as predicted by spectroscopic and computational studies. This work shows the great potential for time-resolved serial femtosecond crystallography for investigation of catalytic processes in biomolecules.
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| 10. |
- Laksmono, Hartawan, et al.
(författare)
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Anomalous Behavior of the Homogeneous Ice Nucleation Rate in No-Man's Land
- 2015
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Ingår i: Journal of Physical Chemistry Letters. - : American Chemical Society (ACS). - 1948-7185 .- 1948-7185. ; 6:14, s. 2826-2832
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Tidskriftsartikel (refereegranskat)abstract
- We present an analysis of ice nucleation kinetics from near-ambient pressure water as temperature decreases below the homogeneous limit T-H by cooling micrometer-sized droplets (microdroplets) evaporatively at 10(3)-10(4) K/s and probing the structure ultrafast using femtosecond pulses from the Linac Coherent Light Source (LCLS) free-electron X-ray laser. Below 232 K, we observed a slower nucleation rate increase with decreasing temperature than anticipated from previous measurements, which we suggest is due to the rapid decrease in water's diffusivity. This is consistent with earlier findings that microdroplets do not crystallize at <227 K, but vitrify at cooling rates of 10(6)-10(7) K/s. We also hypothesize that the slower increase in the nucleation rate is connected with the proposed fragile-to-strong transition anomaly in water.
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