RADDOSE-XFEL: femtosecond time-resolved dose estimates for macromolecular X-ray free-electron laser experiments

2020 ◽  
Vol 53 (2) ◽  
pp. 549-560 ◽  
Author(s):  
Joshua L. Dickerson ◽  
Patrick T. N. McCubbin ◽  
Elspeth F. Garman
Keyword(s):  
Data Collection ◽  
Radiation Damage ◽  
Free Electron ◽  
Absorbed Dose ◽  
Limiting Factor ◽  
Time Resolved ◽  
X Ray ◽  
Laser Experiments ◽  
Beam Parameters ◽  
Diffraction Patterns

For macromolecular structure determination at synchrotron sources, radiation damage remains a major limiting factor. Estimation of the absorbed dose (J kg−1) during data collection at these sources by programs such as RADDOSE-3D has allowed direct comparison of radiation damage between experiments carried out with different samples and beam parameters. This has enabled prediction of roughly when radiation damage will manifest so it can potentially be avoided. X-ray free-electron lasers (XFELs), which produce intense X-ray pulses only a few femtoseconds in duration, can be used to generate diffraction patterns before most of the radiation damage processes have occurred and hence hypothetically they enable the determination of damage-free atomic resolution structures. In spite of this, several experimental and theoretical studies have suggested that structures from XFELs are not always free of radiation damage. There are currently no freely available programs designed to calculate the dose absorbed during XFEL data collection. This article presents an extension to RADDOSE-3D called RADDOSE-XFEL, which calculates the time-resolved dose during XFEL experiments. It is anticipated that RADDOSE-XFEL could be used to facilitate the study of radiation damage at XFELs and ultimately be used prior to data collection so that experimenters can plan their experiments to avoid radiation damage manifesting in their structures.

RADDOSE-3D: time- and space-resolved modelling of dose in macromolecular crystallography

2013 ◽  
Vol 46 (4) ◽  
pp. 1225-1230 ◽  
Author(s):  
Oliver B. Zeldin ◽  
Markus Gerstel ◽  
Elspeth F. Garman
Keyword(s):  
Data Collection ◽  
Radiation Damage ◽  
Diffraction Experiment ◽  
Macromolecular Crystallography ◽  
X Ray Diffraction ◽  
Time Resolved ◽  
X Ray ◽  
Online Methods ◽  
Crystal Volume ◽  
Number Of Iterations

RADDOSE-3D allows the macroscopic modelling of an X-ray diffraction experiment for the purpose of better predicting radiation-damage progression. The distribution of dose within the crystal volume is calculated for a number of iterations in small angular steps across one or more data collection wedges, providing a time-resolved picture of the dose state of the crystal. The code is highly modular so that future contributions from the community can be easily integrated into it, in particular to incorporate online methods for determining the shape of macromolecular crystals and better protocols for imaging real experimental X-ray beam profiles.

scholarly journals Pump-Probe Time-Resolved Serial Femtosecond Crystallography at SACLA: Current Status and Data Collection Strategies

2019 ◽  
Vol 9 (24) ◽  
pp. 5505 ◽  
Author(s):  
Eriko Nango ◽  
Minoru Kubo ◽  
Kensuke Tono ◽  
So Iwata
Keyword(s):  
Data Collection ◽  
Free Electron ◽  
Free Electron Laser ◽  
Current Status ◽  
Optical Pump ◽  
Time Resolved ◽  
Pump Probe ◽  
X Ray ◽  
Collection Strategies ◽  
Serial Femtosecond Crystallography

Structural information on protein dynamics is a critical factor in fully understanding the protein functions. Pump-probe time-resolved serial femtosecond crystallography (TR-SFX) is a recently established technique for visualizing the structural changes or reactions in proteins that are at work with high spatial and temporal resolution. In the pump-probe method, protein microcrystals are continuously delivered from an injector and exposed to an X-ray free-electron laser (XFEL) pulse after a trigger to initiate a reaction, such as light, chemicals, temperature, and electric field, which affords the structural snapshots of intermediates that occur in the protein. We are in the process of developing the device and techniques for pump-probe TR-SFX while using XFEL produced at SPring-8 Angstrom Compact Free-Electron Laser (SACLA). In this paper, we described our current development details and data collection strategies for the optical pump X-ray probe TR-SFX experiment at SACLA and then reported the techniques of in crystallo TR spectroscopy, which is useful in clarifying the nature of reaction that takes place in crystals in advance.

SAXS Instrumentation

2018 ◽  
Author(s):  
Eaton E. Lattman ◽  
Thomas D. Grant ◽  
Edward H. Snell
Keyword(s):  
Data Collection ◽  
Free Electron ◽  
Free Electron Laser ◽  
Sample Volume ◽  
Time Resolved ◽  
Sample Handling ◽  
X Ray ◽  
Efficient Data ◽  
Laser Sources ◽  
Efficient Data Collection

SAXS instrumentation is available for the laboratory, at the synchrotron, and at X-ray free electron laser sources. This chapter deals with SAXS instrumentation. It covers laboratory systems, synchrotron beamlines and newer sources. Multiple synchrotron facilities have SAXS beamlines and the ability to perform SAXS studies on free electron laser sources is growing. Sample handling has adopted automation and in some cases microfluidics to reduce sample volume requirements. Sensitive detectors efficient data collection software and rapid analysis allow real time decisions to be made during data collection. Specialized apparatus enables time resolved studies. Each component is described.

scholarly journals Radiation damage in protein crystallography at X-ray free-electron lasers

2019 ◽  
Vol 75 (2) ◽  
pp. 211-218 ◽  
Author(s):  
Karol Nass
Keyword(s):  
Radiation Damage ◽  
Free Electron ◽  
Heavy Elements ◽  
Limiting Factor ◽  
Theoretical Studies ◽  
Free Electron Lasers ◽  
X Ray ◽  
Protein Molecules ◽  
Damage Processes ◽  
Experimental And Theoretical Studies

Radiation damage is still the most limiting factor in obtaining high-resolution structures of macromolecules in crystallographic experiments at synchrotrons. With the advent of X-ray free-electron lasers (XFELs) that produce ultrashort and highly intense X-ray pulses, it became possible to outrun most of the radiation-damage processes occurring in the sample during exposure to XFEL radiation. Although this is generally the case, several experimental and theoretical studies have indicated that structures from XFELs may not always be radiation-damage free. This is especially true when higher intensity pulses are used and protein molecules that contain heavy elements in their structures are studied. Here, the radiation-damage mechanisms that occur in samples exposed to XFEL pulses are summarized, results that show indications of radiation damage are reviewed and methods that can partially overcome it are discussed.

scholarly journals Radiation damage to biological samples: still a pertinent issue

2021 ◽  
Vol 28 (5) ◽  
Author(s):  
Elspeth F. Garman ◽  
Martin Weik
Keyword(s):  
Data Collection ◽  
Radiation Damage ◽  
Biological Samples ◽  
Imaging Techniques ◽  
Relevant Literature ◽  
Absorbed Dose ◽  
Oh Radicals ◽  
X Rays ◽  
Special Issue ◽  
X Ray

An understanding of radiation damage effects suffered by biological samples during structural analysis using both X-rays and electrons is pivotal to obtain reliable molecular models of imaged molecules. This special issue on radiation damage contains six papers reporting analyses of damage from a range of biophysical imaging techniques. For X-ray diffraction, an in-depth study of multi-crystal small-wedge data collection single-wavelength anomalous dispersion phasing protocols is presented, concluding that an absorbed dose of 5 MGy per crystal was optimal to allow reliable phasing. For small-angle X-ray scattering, experiments are reported that evaluate the efficacy of three radical scavengers using a protein designed to give a clear signature of damage in the form of a large conformational change upon the breakage of a disulfide bond. The use of X-rays to induce OH radicals from the radiolysis of water for X-ray footprinting are covered in two papers. In the first, new developments and the data collection pipeline at the NSLS-II high-throughput dedicated synchrotron beamline are described, and, in the second, the X-ray induced changes in three different proteins under aerobic and low-oxygen conditions are investigated and correlated with the absorbed dose. Studies in XFEL science are represented by a report on simulations of ultrafast dynamics in protic ionic liquids, and, lastly, a broad coverage of possible methods for dose efficiency improvement in modalities using electrons is presented. These papers, as well as a brief synopsis of some other relevant literature published since the last Journal of Synchrotron Radiation Special Issue on Radiation Damage in 2019, are summarized below.

scholarly journals MicroED data collection and processing

2015 ◽  
Vol 71 (4) ◽  
pp. 353-360 ◽  
Author(s):  
Johan Hattne ◽  
Francis E. Reyes ◽  
Brent L. Nannenga ◽  
Dan Shi ◽  
M. Jason de la Cruz ◽  
...  
Keyword(s):  
Data Collection ◽  
Three Dimensional ◽  
Limiting Factor ◽  
Structural Studies ◽  
X Ray ◽  
X Ray Crystallography ◽  
Atomic Structures ◽  
Single Well ◽  
Diffraction Patterns ◽  
Three Dimensional Models

MicroED, a method at the intersection of X-ray crystallography and electron cryo-microscopy, has rapidly progressed by exploiting advances in both fields and has already been successfully employed to determine the atomic structures of several proteins from sub-micron-sized, three-dimensional crystals. A major limiting factor in X-ray crystallography is the requirement for large and well ordered crystals. By permitting electron diffraction patterns to be collected from much smaller crystals, or even single well ordered domains of large crystals composed of several small mosaic blocks, MicroED has the potential to overcome the limiting size requirement and enable structural studies on difficult-to-crystallize samples. This communication details the steps for sample preparation, data collection and reduction necessary to obtain refined, high-resolution, three-dimensional models by MicroED, and presents some of its unique challenges.

scholarly journals Analysis of Multi-Hit Crystals in Serial Synchrotron Crystallography Experiments Using High-Viscosity Injectors

Crystals ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 49
Author(s):  
Marjan Hadian-Jazi ◽  
Peter Berntsen ◽  
Hugh Marman ◽  
Brian Abbey ◽  
Connie Darmanin
Keyword(s):  
Radiation Damage ◽  
Absorbed Dose ◽  
High Viscosity ◽  
Interaction Region ◽  
Frame Rate ◽  
Silicone Grease ◽  
Time Resolved ◽  
X Ray ◽  
Sample Flow Rate ◽  
Free Data

Serial Synchrotron Crystallography (SSX) is rapidly emerging as a promising technique for collecting data for time-resolved structural studies or for performing room temperature micro-crystallography measurements using micro-focused beamlines. SSX is often performed using high frame rate detectors in combination with continuous sample scanning or high-viscosity or liquid jet injectors. When performed using ultra-bright X-ray Free Electron Laser (XFEL) sources serial crystallography typically involves a process known as ’diffract-and-destroy’ where each crystal is measured just once before it is destroyed by the intense XFEL pulse. In SSX, however, particularly when using high-viscosity injectors (HVIs) such as Lipidico, the crystal can be intercepted multiple times by the X-ray beam prior to exiting the interaction region. This has a number of important consequences for SSX including whether these multiple-hits can be incorporated into the data analysis or whether they need to be excluded due to the potential impact of radiation damage. Here, we investigate the occurrence and characteristics of multiple hits on single crystals using SSX with lipidico. SSX data are collected from crystals as they tumble within a high viscous stream of silicone grease flowing through a micro-focused X-ray beam. We confirmed that, using the Eiger 16M, we are able to collect up to 42 frames of data from the same single crystal prior to it leaving the X-ray interaction region. The frequency and occurrence of multiple hits may be controlled by varying the sample flow rate and X-ray beam size. Calculations of the absorbed dose confirm that these crystals are likely to undergo radiation damage but that nonetheless incorporating multiple hits into damage-free data should lead to a significant reduction in the number of crystals required for structural analysis when compared to just looking at a single diffraction pattern from each crystal.

Factors in Photographic Emulsions for Low Dose Electron Crystallography

1980 ◽  
Vol 38 ◽  
pp. 230-231
Author(s):  
T.W. Jeng ◽  
W. Chiu
Keyword(s):  
Radiation Damage ◽  
Low Dose ◽  
Damage Effect ◽  
Photographic Emulsion ◽  
Electron Crystallography ◽  
Electron Microscopic ◽  
X Ray ◽  
Signal Contrast ◽  
Diffraction Patterns ◽  
Photographic Emulsions

With the advances in preparing biological materials in a thin and highly ordered form, and in maintaining them hydrated under vacuum, electron crystallography has become an important tool for biological structure investigation at high resolution (1,2). However, the electron radiation damage would limit the capability of recording reflections with low intensities in an electron diffraction pattern. It has been demonstrated that the use of a low temperature stage can reduce the radiation damage effect and that one can expose the specimen with a higher dose in order to increase the signal contrast (3). A further improvement can be made by selecting a proper photographic emulsion. The primary factors in evaluating the suitability of photographic emulsion for recording low dose diffraction patterns are speed, fog level, electron response at low electron exposure, linearity, and usable range of exposure. We have compared these factors with three photographic emulsions including Kodak electron microscopic plate (EMP), Industrex AA x-ray film (AA x-ray) and Kodak nuclear track film (NTB3).

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