| 1. |
- Khakurel, Krishna P., et al.
(author)
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Kilohertz Macromolecular Crystallography Using an EIGER Detector at Low X-ray Fluxes
- 2020
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In: Crystals. - : MDPI. - 2073-4352. ; 10:12
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Journal article (peer-reviewed)abstract
- Time-resolved in-house macromolecular crystallography is primarily limited by the capabilities of the in-house X-ray sources. These sources can only provide a time-averaged structure of the macromolecules. A significant effort has been made in the development of in-house laser-driven ultrafast X-ray sources, with one of the goals as realizing the visualization of the structural dynamics of macromolecules at a very short timescale within the laboratory-scale infrastructure. Most of such in-house ultrafast X-ray sources are operated at high repetition rates and usually deliver very low flux. Therefore, the necessity of a detector that can operate at the repetition rate of the laser and perform extremely well under low flux conditions is essential. Here, we present experimental results demonstrating the usability of the hybrid-pixel detectors, such as Eiger X 1M, and provide experimental proof that they can be successfully operated to collect macromolecular crystallographic data up to a detector frame rate of 3 kHz from synchrotron sources. Our results also show that the data reduction and structural analysis are successful at such high frame rates and fluxes as low as 10(8) photons/s, which is comparable to the values expected from a typical laser-driven X-ray source.
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| 2. |
- Oelze, Tim, et al.
(author)
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THz streak camera performance for single-shot characterization of XUV pulses with complex temporal structures
- 2020
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In: Optics Express. - : OPTICAL SOC AMER. - 1094-4087. ; 28:14, s. 20686-20703
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Journal article (peer-reviewed)abstract
- The THz-field-driven streak camera has proven to be a powerful diagnostic-technique that enables the shot-to-shot characterization of the duration and the arrival time jitter of free electron laser (FEL) pulses. Here we investigate the performance of three computational approaches capable to determine the duration of FEL pulses with complex temporal structures from single-shot measurements of up to three simultaneously recorded spectra. We use numerically simulated FEL pulses in order to validate the accuracy of the pulse length retrieval in average as well as in a single-shot mode. We discuss requirements for the THz field strength in order to achieve reliable results and compare our numerical study with the analysis of experimental data that were obtained at the FEL in Hamburg - FLASH. (C) 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreement
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| 3. |
- Polovinkin, Vitaly, et al.
(author)
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Demonstration of electron diffraction from membrane protein crystals grown in a lipidic mesophase after lamella preparation by focused ion beam milling at cryogenic temperatures
- 2020
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In: Journal of applied crystallography. - 0021-8898 .- 1600-5767. ; 53, s. 1416-1424
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Journal article (peer-reviewed)abstract
- Electron crystallography of sub-micrometre-sized 3D protein crystals has emerged recently as a valuable field of structural biology. In meso crystallization methods, utilizing lipidic mesophases, particularly lipidic cubic phases (LCPs), can produce high-quality 3D crystals of membrane proteins (MPs). A major step towards realizing 3D electron crystallography of MP crystals, grown in meso, is to demonstrate electron diffraction from such crystals. The first task is to remove the viscous and sticky lipidic matrix that surrounds the crystals without damaging the crystals. Additionally, the crystals have to be thin enough to let electrons traverse them without significant multiple scattering. In the present work, the concept that focused ion beam milling at cryogenic temperatures (cryo-FIB milling) can be used to remove excess host lipidic mesophase matrix is experimentally verified, and then the crystals are thinned to a thickness suitable for electron diffraction. In this study, bacteriorhodopsin (BR) crystals grown in a lipidic cubic mesophase of monoolein were used as a model system. LCP from a part of a hexagon-shaped plate-like BR crystal (∼10 µm in thickness and ∼70 µm in the longest dimension), which was flash-frozen in liquid nitrogen, was milled away with a gallium FIB under cryogenic conditions, and a part of the crystal itself was thinned into a ∼210 nm-thick lamella with the ion beam. The frozen sample was then transferred into an electron cryo-microscope, and a nanovolume of ∼1400 × 1400 × 210 nm of the BR lamella was exposed to 200 kV electrons at a fluence of ∼0.06 e Å−2. The resulting electron diffraction peaks were detected beyond 2.7 Å resolution (with an average peak height to background ratio of >2) by a CMOS-based Ceta 16M camera. The results demonstrate that cryo-FIB milling produces high-quality lamellae from crystals grown in lipidic mesophases and pave the way for 3D electron crystallography on crystals grown or embedded in highly viscous media.
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