Time-Resolved Serial Femtosecond Crystallography at the European X-ray Free Electron Laser

2019 ◽  
Author(s):  
Marius Schmidt ◽  
Suraj Pandey ◽  
Adrian Mancuso ◽  
Richard Bean
Keyword(s):  
Free Electron ◽  
Free Electron Laser ◽  
Crystallographic Data ◽  
Time Resolved ◽  
X Ray ◽  
Serial Femtosecond Crystallography

Abstract This protocol introduces step by step into the collection of time resolved crystallographic data and their analysis at the European Free Electron Laser.

scholarly journals Time-resolved serial femtosecond crystallography studies at an X-ray free electron laser reveals structural changes in bacteriorhodopsin

2018 ◽  
Vol 1859 ◽  
pp. e28
Author(s):  
Richard Neutze ◽  
Eriko Nango ◽  
Tomoyuki Tanaka ◽  
Takanori Nakane ◽  
So Iwata ◽  
...  
Keyword(s):  
Free Electron ◽  
Free Electron Laser ◽  
Structural Changes ◽  
Time Resolved ◽  
X Ray ◽  
Serial Femtosecond Crystallography

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.

scholarly journals X-ray free-electron laser studies reveal correlated motion during isopenicillin N synthase catalysis

2021 ◽  
Vol 7 (34) ◽  
pp. eabh0250
Author(s):  
Patrick Rabe ◽  
Jos J. A. G. Kamps ◽  
Kyle D. Sutherlin ◽  
James D. S. Linyard ◽  
Pierre Aller ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Free Electron ◽  
Free Electron Laser ◽  
Enzyme Dynamics ◽  
Time Resolved ◽  
X Ray ◽  
Unique Reaction ◽  
Solution Studies ◽  
Correlated Motion ◽  
Serial Femtosecond Crystallography

Isopenicillin N synthase (IPNS) catalyzes the unique reaction of l-δ-(α-aminoadipoyl)-l-cysteinyl-d-valine (ACV) with dioxygen giving isopenicillin N (IPN), the precursor of all natural penicillins and cephalosporins. X-ray free-electron laser studies including time-resolved crystallography and emission spectroscopy reveal how reaction of IPNS:Fe(II):ACV with dioxygen to yield an Fe(III) superoxide causes differences in active site volume and unexpected conformational changes that propagate to structurally remote regions. Combined with solution studies, the results reveal the importance of protein dynamics in regulating intermediate conformations during conversion of ACV to IPN. The results have implications for catalysis by multiple IPNS-related oxygenases, including those involved in the human hypoxic response, and highlight the power of serial femtosecond crystallography to provide insight into long-range enzyme dynamics during reactions presently impossible for nonprotein catalysts.

Novel opportunities for time-resolved absorption spectroscopy at the X-ray free electron laser

2010 ◽  
Vol 12 (21) ◽  
pp. 5647 ◽  
Author(s):  
B. D. Patterson ◽  
R. Abela
Keyword(s):  
Absorption Spectroscopy ◽  
Free Electron ◽  
Free Electron Laser ◽  
Time Resolved ◽  
X Ray

scholarly journals Liquid sample delivery techniques for serial femtosecond crystallography

2014 ◽  
Vol 369 (1647) ◽  
pp. 20130337 ◽  
Author(s):  
Uwe Weierstall
Keyword(s):  
Flow Rate ◽  
Free Electron ◽  
Repetition Rate ◽  
Free Electron Laser ◽  
Room Temperature ◽  
Liquid Flow Rate ◽  
Free Electron Lasers ◽  
Liquid Sample ◽  
X Ray ◽  
Serial Femtosecond Crystallography

X-ray free-electron lasers overcome the problem of radiation damage in protein crystallography and allow structure determination from micro- and nanocrystals at room temperature. To ensure that consecutive X-ray pulses do not probe previously exposed crystals, the sample needs to be replaced with the X-ray repetition rate, which ranges from 120 Hz at warm linac-based free-electron lasers to 1 MHz at superconducting linacs. Liquid injectors are therefore an essential part of a serial femtosecond crystallography experiment at an X-ray free-electron laser. Here, we compare different techniques of injecting microcrystals in solution into the pulsed X-ray beam in vacuum. Sample waste due to mismatch of the liquid flow rate to the X-ray repetition rate can be addressed through various techniques.

scholarly journals Two mirror X-ray pulse split and delay instrument for femtosecond time resolved investigations at the LCLS free electron laser facility

2016 ◽  
Vol 24 (11) ◽  
pp. 11768 ◽  
Author(s):  
Nora Berrah ◽  
Li Fang ◽  
Brendan F Murphy ◽  
Edwin Kukk ◽  
Timur Y. Osipov ◽  
...  
Keyword(s):  
Free Electron ◽  
Free Electron Laser ◽  
Time Resolved ◽  
X Ray ◽  
Laser Facility

scholarly journals Direct autocorrelation of soft-x-ray free-electron-laser pulses by time-resolved two-photon double ionization of He

2009 ◽  
Vol 80 (2) ◽  
Author(s):  
R. Mitzner ◽  
A. A. Sorokin ◽  
B. Siemer ◽  
S. Roling ◽  
M. Rutkowski ◽  
...  
Keyword(s):  
Free Electron ◽  
Free Electron Laser ◽  
Laser Pulses ◽  
Double Ionization ◽  
Time Resolved ◽  
X Ray ◽  
Two Photon

scholarly journals Pink-beam serial femtosecond crystallography for accurate structure-factor determination at an X-ray free-electron laser

IUCrJ ◽  
2021 ◽  
Vol 8 (6) ◽  
Author(s):  
Karol Nass ◽  
Camila Bacellar ◽  
Claudio Cirelli ◽  
Florian Dworkowski ◽  
Yaroslav Gevorkov ◽  
...  
Keyword(s):  
Data Quality ◽  
Structure Determination ◽  
Structure Factor ◽  
Free Electron ◽  
Free Electron Laser ◽  
De Novo ◽  
Similar Data ◽  
X Ray ◽  
Redundant Data ◽  
Serial Femtosecond Crystallography

Serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs) enables essentially radiation-damage-free macromolecular structure determination using microcrystals that are too small for synchrotron studies. However, SFX experiments often require large amounts of sample in order to collect highly redundant data where some of the many stochastic errors can be averaged out to determine accurate structure-factor amplitudes. In this work, the capability of the Swiss X-ray free-electron laser (SwissFEL) was used to generate large-bandwidth X-ray pulses [Δλ/λ = 2.2% full width at half-maximum (FWHM)], which were applied in SFX with the aim of improving the partiality of Bragg spots and thus decreasing sample consumption while maintaining the data quality. Sensitive data-quality indicators such as anomalous signal from native thaumatin micro-crystals and de novo phasing results were used to quantify the benefits of using pink X-ray pulses to obtain accurate structure-factor amplitudes. Compared with data measured using the same setup but using X-ray pulses with typical quasi-monochromatic XFEL bandwidth (Δλ/λ = 0.17% FWHM), up to fourfold reduction in the number of indexed diffraction patterns required to obtain similar data quality was achieved. This novel approach, pink-beam SFX, facilitates the yet underutilized de novo structure determination of challenging proteins at XFELs, thereby opening the door to more scientific breakthroughs.

scholarly journals Batch crystallization of rhodopsin for structural dynamics using an X-ray free-electron laser

2015 ◽  
Vol 71 (7) ◽  
pp. 856-860 ◽  
Author(s):  
Wenting Wu ◽  
Przemyslaw Nogly ◽  
Jan Rheinberger ◽  
Leonhard M. Kick ◽  
Cornelius Gati ◽  
...  
Keyword(s):  
Free Electron ◽  
Free Electron Laser ◽  
Second Harmonic ◽  
Night Vision ◽  
Salt Crystallization ◽  
Time Resolved ◽  
Batch Crystallization ◽  
X Ray ◽  
Vapour Diffusion ◽  
Serial Crystallography

Rhodopsin is a membrane protein from the G protein-coupled receptor family. Together with its ligand retinal, it forms the visual pigment responsible for night vision. In order to perform ultrafast dynamics studies, a time-resolved serial femtosecond crystallography method is required owing to the nonreversible activation of rhodopsin. In such an approach, microcrystals in suspension are delivered into the X-ray pulses of an X-ray free-electron laser (XFEL) after a precise photoactivation delay. Here, a millilitre batch production of high-density microcrystals was developed by four methodical conversion steps starting from known vapour-diffusion crystallization protocols: (i) screening the low-salt crystallization conditions preferred for serial crystallography by vapour diffusion, (ii) optimization of batch crystallization, (iii) testing the crystal size and quality using second-harmonic generation (SHG) imaging and X-ray powder diffraction and (iv) production of millilitres of rhodopsin crystal suspension in batches for serial crystallography tests; these crystals diffracted at an XFEL at the Linac Coherent Light Source using a liquid-jet setup.

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