| 1. |
- Ayyer, Kartik, et al.
(författare)
-
3D diffractive imaging of nanoparticle ensembles using an x-ray laser
- 2021
-
Ingår i: Optica. - : Optical Society of America. - 2334-2536. ; 8:1, s. 15-23
-
Tidskriftsartikel (refereegranskat)abstract
- Single particle imaging at x-ray free electron lasers (XFELs) has the potential to determine the structure and dynamics of single biomolecules at room temperature. Two major hurdles have prevented this potential from being reached, namely, the collection of sufficient high-quality diffraction patterns and robust computational purification to overcome structural heterogeneity. We report the breaking of both of these barriers using gold nanoparticle test samples, recording around 10 million diffraction patterns at the European XFEL and structurally and orientationally sorting the patterns to obtain better than 3-nm-resolution 3D reconstructions for each of four samples. With these new developments, integrating advancements in x-ray sources, fast-framing detectors, efficient sample delivery, and data analysis algorithms, we illuminate the path towards sub-nano meter biomolecular imaging. The methods developed here can also be extended to characterize ensembles that are inherently diverse to obtain their full structural landscape. Published by The Optical Society under the terms of the Creative Commons Attribution 4.0 License.
|
|
| 2. |
- Johansson, Linda C, 1983, et al.
(författare)
-
Lipidic phase membrane protein serial femtosecond crystallography.
- 2012
-
Ingår i: Nature methods. - : Springer Science and Business Media LLC. - 1548-7105 .- 1548-7091. ; 9:3, s. 263-265
-
Tidskriftsartikel (refereegranskat)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.
|
|
| 3. |
- Sobolev, Egor, et al.
(författare)
-
Megahertz single-particle imaging at the European XFEL
- 2020
-
Ingår i: Communications Physics. - : Springer Science and Business Media LLC. - 2399-3650. ; 3:1
-
Tidskriftsartikel (refereegranskat)abstract
- The emergence of high repetition-rate X-ray free-electron lasers (XFELs) powered by superconducting accelerator technology enables the measurement of significantly more experimental data per day than was previously possible. The European XFEL is expected to provide 27,000 pulses per second, over two orders of magnitude more than any other XFEL. The increased pulse rate is a key enabling factor for single-particle X-ray diffractive imaging, which relies on averaging the weak diffraction signal from single biological particles. Taking full advantage of this new capability requires that all experimental steps, from sample preparation and delivery to the acquisition of diffraction patterns, are compatible with the increased pulse repetition rate. Here, we show that single-particle imaging can be performed using X-ray pulses at megahertz repetition rates. The results obtained pave the way towards exploiting high repetition-rate X-ray free-electron lasers for single-particle imaging at their full repetition rate.
|
|
| 4. |
|
|
| 5. |
- Aquila, Andrew, et al.
(författare)
-
Time-resolved protein nanocrystallography using an X-ray free-electron laser
- 2012
-
Ingår i: Optics Express. - 1094-4087. ; 20:3, s. 2706-2716
-
Tidskriftsartikel (refereegranskat)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.
|
|
| 6. |
- Chapman, Henry N, et al.
(författare)
-
Femtosecond X-ray protein nanocrystallography.
- 2011
-
Ingår i: Nature. - : Springer Science and Business Media LLC. - 1476-4687 .- 0028-0836. ; 470:7332, s. 73-7
-
Tidskriftsartikel (refereegranskat)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 (∼200nm to 2μm 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.
|
|
| 7. |
- Ekeberg, Tomas, 1983-, et al.
(författare)
-
Observation of a single protein by ultrafast X-ray diffraction
- 2024
-
Ingår i: Light. - : Springer Nature. - 2095-5545 .- 2047-7538. ; 13:1
-
Tidskriftsartikel (refereegranskat)abstract
- The idea of using ultrashort X-ray pulses to obtain images of single proteins frozen in time has fascinated and inspired many. It was one of the arguments for building X-ray free-electron lasers. According to theory, the extremely intense pulses provide sufficient signal to dispense with using crystals as an amplifier, and the ultrashort pulse duration permits capturing the diffraction data before the sample inevitably explodes. This was first demonstrated on biological samples a decade ago on the giant mimivirus. Since then, a large collaboration has been pushing the limit of the smallest sample that can be imaged. The ability to capture snapshots on the timescale of atomic vibrations, while keeping the sample at room temperature, may allow probing the entire conformational phase space of macromolecules. Here we show the first observation of an X-ray diffraction pattern from a single protein, that of Escherichia coli GroEL which at 14 nm in diameter is the smallest biological sample ever imaged by X-rays, and demonstrate that the concept of diffraction before destruction extends to single proteins. From the pattern, it is possible to determine the approximate orientation of the protein. Our experiment demonstrates the feasibility of ultrafast imaging of single proteins, opening the way to single-molecule time-resolved studies on the femtosecond timescale.
|
|
| 8. |
- Kassemeyer, Stephan, et al.
(författare)
-
Femtosecond free-electron laser x-ray diffraction data sets for algorithm development
- 2012
-
Ingår i: Optics Express. - 1094-4087. ; 20:4, s. 4149-4158
-
Tidskriftsartikel (refereegranskat)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.
|
|
| 9. |
- Kirian, Richard A., et al.
(författare)
-
Structure-factor analysis of femtosecond micro-diffraction patterns from protein nanocrystals
- 2011
-
Ingår i: Acta Crystallographica Section A. - 0108-7673 .- 1600-5724. ; 67:2, s. 131-140
-
Tidskriftsartikel (refereegranskat)abstract
- A complete set of structure factors has been extracted from hundreds of thousands of femtosecond single-shot X-ray microdiffraction patterns taken from randomly oriented nanocrystals. The method of Monte Carlo integration over crystallite size and orientation was applied to experimental data from Photosystem I nanocrystals. This arrives at structure factors from many partial reflections without prior knowledge of the particle-size distribution. The data were collected at the Linac Coherent Light Source (the first hard-X-ray laser user facility), to which was fitted a hydrated protein nanocrystal injector jet, according to the method of serial crystallography. The data are single 'still' diffraction snapshots, each from a different nanocrystal with sizes ranging between 100 nm and 2 mu m, so the angular width of Bragg peaks was dominated by crystal-size effects. These results were compared with single-crystal data recorded from large crystals of Photosystem I at the Advanced Light Source and the quality of the data was found to be similar. The implications for improving the efficiency of data collection by allowing the use of very small crystals, for radiation-damage reduction and for time-resolved diffraction studies at room temperature are discussed.
|
|
| 10. |
- Koopmann, Rudolf, et al.
(författare)
-
In vivo protein crystallization opens new routes in structural biology
- 2012
-
Ingår i: Nature Methods. - : Springer Science and Business Media LLC. - 1548-7091 .- 1548-7105. ; 9:3, s. 259-262
-
Tidskriftsartikel (refereegranskat)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.
|
|
