Macromolecular nanocrystal structural analysis with electron and X-rays: A comparative review

Crystallography has long been the unrivaled method that can provide the atomistic structural models of macromolecules, using either X-rays or electrons as probes. The methodology has gone through several revolutionary periods, driven by the development of new sources, detectors, and other instrumentation. Novel sources of both X-ray and electrons are constantly emerging. The increase in brightness of these sources, complemented by the advanced detection techniques, has relaxed the traditionally strict need for large, high quality, crystals. Recent reports suggest high-quality diffraction datasets from crystals as small as a few hundreds of nanometers can be routinely obtained. This has resulted in the genesis of a new field of macromolecular nanocrystal crystallography. Here we will make a brief comparative review of this growing field focusing on the use of X-rays and electrons sources

Progress and Poverty—1965 Version

The first hard X-ray laser, the Linac Coherent Light Source (LCLS), produces 120 shots per second. Particles injected into the X-ray beam are hit randomly and in unknown orientations by the extremely intense X-ray pulses, where the femtosecond-duration X-ray pulses diffract from the sample before the particle structure is significantly changed even though the sample is ultimately destroyed by the deposited X-ray energy. Single particle X-ray diffraction experiments generate data at the FEL repetition rate, resulting in more than 400,000 detector readouts in an hour, the data stream during an experiment contains blank frames mixed with hits on single particles, clusters and contaminants. The diffraction signal is generally weak and it is superimposed on a low but continually fluctuating background signal, originating from photon noise in the beam line and electronic noise from the detector. Meanwhile, explosion of the sample creates fragments with a characteristic signature. Here, we describe methods based on rapid image analysis combined with ion Time-of-Flight (ToF) spectroscopy of the fragments to achieve an efficient, automated and unsupervised sorting of diffraction data. The studies described here form a basis for the development of real-time frame rejection methods, e. g. for the European XFEL, which is expected to produce 100 million pulses per hour. (C)2014 Optical Society of Americ

Phonon spectra of pure and acceptor doped BaZrO3 investigated with visible and UV Raman spectroscopy

We report results from visible and UV Raman spectroscopy studies of the phonon spectra of a polycrystalline sample of the prototypical perovskite type oxide BaZrO3 and a 500 nm thick film of its Y-doped, proton conducting, counterpart BaZr0.8Y0.2O2.9. Analysis of the Raman spectra measured using different excitation energies (between 3.44 eV and 5.17 eV) reveals the activation of strong resonance Raman effects involving all lattice vibrational modes. Specifically, two characteristic energies were identified for BaZrO3, one around 5 eV and one at higher energy, respectively, and one for BaZr0.8Y0.2O2.9, above 5 eV. Apart from the large difference in spectral intensity between the non-resonant and resonant conditions, the spectra are overall similar to each other, suggesting that the vibrational spectra of the perovskites are stable when investigated using an UV laser as excitation source. These results encourage further use of UV Raman spectroscopy as a novel approach for the study of lattice vibrational dynamics and local structure in proton conducting perovskites, and open up for, e.g., time-resolved experiments on thin films targeted at understanding the role of lattice vibrations in proton transport in these kinds of materials

Ultrafast multi-electron dynamics studied with THz-field streaking

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A statistical approach to detect protein complexes at X-ray free electron laser facilities

The Flash X-ray Imaging (FXI) technique, under development at X-ray free electron lasers (XFEL), aims to achieve structure determination based on diffraction from individual macromolecular complexes. We report an FXI study on the first protein complex-RNA polymerase II-ever injected at an XFEL. A successful 3D reconstruction requires a high number of observations of the sample in various orientations. The measured diffraction signal for many shots can be comparable to background. Here we present a robust and highly sensitive hit-identification method based on automated modeling of beamline background through photon statistics. It can operate at controlled false positive hit-rate of 3 x10(-5). We demonstrate its power in determining particle hits and validate our findings against an independent hit-identification approach based on ion time-of-flight spectra. We also validate the advantages of our method over simpler hit-identification schemes via tests on other samples and using computer simulations, showing a doubled hit-identification power

THz streak camera performance for single-shot characterization of XUV pulses with complex temporal structures

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. © 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreemen

Coherent diffraction of single Rice Dwarf virus particles using hard X-rays at the Linac Coherent Light Source

Single particle diffractive imaging data from Rice Dwarf Virus (RDV) were recorded using the Coherent X-ray Imaging (CXI) instrument at the Linac Coherent Light Source (LCLS). RDV was chosen as it is a wellcharacterized model system, useful for proof-of-principle experiments, system optimization and algorithm development. RDV, an icosahedral virus of about 70 nm in diameter, was aerosolized and injected into the approximately 0.1 mu m diameter focused hard X-ray beam at the CXI instrument of LCLS. Diffraction patterns from RDV with signal to 5.9 angstrom ngstrom were recorded. The diffraction data are available through the Coherent X-ray Imaging Data Bank (CXIDB) as a resource for algorithm development, the contents of which are described here.11Ysciescopu

Megahertz single-particle imaging at the European XFEL

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.We acknowledge European XFEL in Schenefeld, Germany, for provision of X-ray free-electron laser beamtime at Scientific Instrument SPB/SFX and would like to thank the instrument group and facility staff for their assistance. We acknowledge the use of the XBI biological sample preparation laboratory, enabled by the XBI User Consortium. The results of the work were obtained using Maxwell computational resources operated at Deutsches Elektronen-Synchrotron (DESY), Hamburg, Germany, and computational resources of MCC NRC “Kurchatov Institute.” This research used resources of the National Synchrotron Light Source II, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Brookhaven National Laboratory under Contract No. DE-SC0012704. We acknowledge the support of funding from: the Swedish Foundation for International Cooperation in Research and Higher Education (STINT); Helmholtz Associations Initiative and Networking Fund and the Russian Science Foundation grant HRSF-0002/18-41-0600; the Russian Science Foundation grant 18-14-00321; European Research Council, “Frontiers in Attosecond X-ray Science: Imaging and Spectroscopy (AXSIS),” ERC-2013-SyG 609920; Fellowship from the Joachim Herz Stiftung; Singapore National Research Foundation Grant number NRF-CRP16-2015-05; Ministry of Education, Science, Research and Sport of the Slovak Republic and by grant APVV-18-0104; the project CZ.02.1.01/0.0/0.0/16_019/0000789 (ADONIS) from European Regional Development Fund, Chalmers Area of Advance; Material Science and the Ministry of Education, Youth and Sports as part of targeted support from the National Programme of Sustainability II; US National Science Foundation (NSF) Science and Technology Center BioXFEL Award 1231306; Helmholtz Initiative and Networking Fund through the Young Investigators Group Program and Deutsche Forschungsgemeinschaft, project B03/SFB755; VR starting grant (2018-03387); FORMAS future research leader (2018-00421); KVA Biosciences 2018 (BS2018-0053); NSF 1231306; German Ministry for Education and Research, BMBF (grant No. 05K2016—Visavix); the Heinrich Pette Institute, Leibniz Institute for Experimental Virology is supported by the Free and Hanseatic City of Hamburg and the Federal Ministry of Health; NSF STC BioXFEL grant 1231306; The National Research Foundation (NRF) of Korea (Grant No. 2017K1A3A7A09016380); the Röntgen-Ångström Cluster; the Swedish Research Council; the Swedish Foundation for Strategic Research. We thank Arwen Pearson for critical reading of the manuscript. Open access funding provided by Uppsala University