| 1. |
- Johansson, Linda C, 1983, et al.
(author)
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Lipidic phase membrane protein serial femtosecond crystallography.
- 2012
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In: Nature methods. - : Springer Science and Business Media LLC. - 1548-7105 .- 1548-7091. ; 9:3, s. 263-265
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Journal article (peer-reviewed)abstract
- X-ray free electron laser (X-FEL)-based serial femtosecond crystallography is an emerging method with potential to rapidly advance the challenging field of membrane protein structural biology. Here we recorded interpretable diffraction data from micrometer-sized lipidic sponge phase crystals of the Blastochloris viridis photosynthetic reaction center delivered into an X-FEL beam using a sponge phase micro-jet.
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| 2. |
- Aquila, Andrew, et al.
(author)
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Time-resolved protein nanocrystallography using an X-ray free-electron laser
- 2012
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In: Optics Express. - 1094-4087. ; 20:3, s. 2706-2716
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Journal article (peer-reviewed)abstract
- We demonstrate the use of an X-ray free electron laser synchronized with an optical pump laser to obtain X-ray diffraction snapshots from the photoactivated states of large membrane protein complexes in the form of nanocrystals flowing in a liquid jet. Light-induced changes of Photosystem I-Ferredoxin co-crystals were observed at time delays of 5 to 10 µs after excitation. The result correlates with the microsecond kinetics of electron transfer from Photosystem I to ferredoxin. The undocking process that follows the electron transfer leads to large rearrangements in the crystals that will terminally lead to the disintegration of the crystals. We describe the experimental setup and obtain the first time-resolved femtosecond serial X-ray crystallography results from an irreversible photo-chemical reaction at the Linac Coherent Light Source. This technique opens the door to time-resolved structural studies of reaction dynamics in biological systems.
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| 3. |
- Chapman, Henry N, et al.
(author)
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Femtosecond X-ray protein nanocrystallography.
- 2011
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In: Nature. - : Springer Science and Business Media LLC. - 1476-4687 .- 0028-0836. ; 470:7332, s. 73-7
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Journal article (peer-reviewed)abstract
- X-ray crystallography provides the vast majority of macromolecular structures, but the success of the method relies on growing crystals of sufficient size. In conventional measurements, the necessary increase in X-ray dose to record data from crystals that are too small leads to extensive damage before a diffraction signal can be recorded. It is particularly challenging to obtain large, well-diffracting crystals of membrane proteins, for which fewer than 300 unique structures have been determined despite their importance in all living cells. Here we present a method for structure determination where single-crystal X-ray diffraction 'snapshots' are collected from a fully hydrated stream of nanocrystals using femtosecond pulses from a hard-X-ray free-electron laser, the Linac Coherent Light Source. We prove this concept with nanocrystals of photosystem I, one of the largest membrane protein complexes. More than 3,000,000 diffraction patterns were collected in this study, and a three-dimensional data set was assembled from individual photosystem I nanocrystals (∼200 nm to 2 μm in size). We mitigate the problem of radiation damage in crystallography by using pulses briefer than the timescale of most damage processes. This offers a new approach to structure determination of macromolecules that do not yield crystals of sufficient size for studies using conventional radiation sources or are particularly sensitive to radiation damage.
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| 4. |
- Kassemeyer, Stephan, et al.
(author)
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Femtosecond free-electron laser x-ray diffraction data sets for algorithm development
- 2012
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In: Optics Express. - 1094-4087. ; 20:4, s. 4149-4158
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Journal article (peer-reviewed)abstract
- We describe femtosecond X-ray diffraction data sets of viruses and nanoparticles collected at the Linac Coherent Light Source. The data establish the first large benchmark data sets for coherent diffraction methods freely available to the public, to bolster the development of algorithms that are essential for developing this novel approach as a useful imaging technique. Applications are 2D reconstructions, orientation classification and finally 3D imaging by assembling 2D patterns into a 3D diffraction volume.
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| 5. |
- Koopmann, Rudolf, et al.
(author)
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In vivo protein crystallization opens new routes in structural biology
- 2012
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In: Nature Methods. - : Springer Science and Business Media LLC. - 1548-7091 .- 1548-7105. ; 9:3, s. 259-262
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Journal article (peer-reviewed)abstract
- Protein crystallization in cells has been observed several times in nature. However, owing to their small size these crystals have not yet been used for X-ray crystallographic analysis. We prepared nano-sized in vivo–grown crystals of Trypanosoma brucei enzymes and applied the emerging method of free-electron laser-based serial femtosecond crystallography to record interpretable diffraction data. This combined approach will open new opportunities in structural systems biology.
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| 6. |
- Lomb, Lukas, et al.
(author)
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Radiation damage in protein serial femtosecond crystallography using an x-ray free-electron laser
- 2011
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In: Physical Review B. Condensed Matter and Materials Physics. - 1098-0121 .- 1550-235X. ; 84:21, s. 214111-1-214111-6
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Journal article (peer-reviewed)abstract
- X-ray free-electron lasers deliver intense femtosecond pulses that promise to yield high resolution diffraction data of nanocrystals before the destruction of the sample by radiation damage. Diffraction intensities of lysozyme nanocrystals collected at the Linac Coherent Light Source using 2 keV photons were used for structure determination by molecular replacement and analyzed for radiation damage as a function of pulse length and fluence. Signatures of radiation damage are observed for pulses as short as 70 fs. Parametric scaling used in conventional crystallography does not account for the observed effects.
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| 7. |
- Seibert, M. Marvin, et al.
(author)
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Single mimivirus particles intercepted and imaged with an X-ray laser
- 2011
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In: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 470:7332, s. 78-81
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Journal article (peer-reviewed)abstract
- X-ray lasers offer new capabilities in understanding the structure of biological systems, complex materials and matter under extreme conditions(1-4). Very short and extremely bright, coherent X-ray pulses can be used to outrun key damage processes and obtain a single diffraction pattern from a large macromolecule, a virus or a cell before the sample explodes and turns into plasma(1). The continuous diffraction pattern of non-crystalline objects permits oversampling and direct phase retrieval(2). Here we show that high-quality diffraction data can be obtained with a single X-ray pulse from a noncrystalline biological sample, a single mimivirus particle, which was injected into the pulsed beam of a hard-X-ray free-electron laser, the Linac Coherent Light Source(5). Calculations indicate that the energy deposited into the virus by the pulse heated the particle to over 100,000 K after the pulse had left the sample. The reconstructed exit wavefront (image) yielded 32-nm full-period resolution in a single exposure and showed no measurable damage. The reconstruction indicates inhomogeneous arrangement of dense material inside the virion. We expect that significantly higher resolutions will be achieved in such experiments with shorter and brighter photon pulses focused to a smaller area. The resolution in such experiments can be further extended for samples available in multiple identical copies.
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| 8. |
- Kuepper, Jochen, et al.
(author)
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X-Ray Diffraction from Isolated and Strongly Aligned Gas-Phase Molecules with a Free-Electron Laser
- 2014
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In: Physical Review Letters. - 0031-9007 .- 1079-7114. ; 112:8, s. 083002-
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Journal article (peer-reviewed)abstract
- We report experimental results on x-ray diffraction of quantum-state-selected and strongly aligned ensembles of the prototypical asymmetric rotor molecule 2,5-diiodobenzonitrile using the Linac Coherent Light Source. The experiments demonstrate first steps toward a new approach to diffractive imaging of distinct structures of individual, isolated gas-phase molecules. We confirm several key ingredients of single molecule diffraction experiments: the abilities to detect and count individual scattered x-ray photons in single shot diffraction data, to deliver state-selected, e.g., structural-isomer-selected, ensembles of molecules to the x-ray interaction volume, and to strongly align the scattering molecules. Our approach, using ultrashort x-ray pulses, is suitable to study ultrafast dynamics of isolated molecules.
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