scholarly journals De novo phasing of protein crystallography data from a free-electron laser

2014 ◽  
Vol 70 (a1) ◽  
pp. C568-C568
Author(s):  
Thomas Barends ◽  
Lutz Foucar ◽  
Sabine Botha ◽  
R. Bruce Doak ◽  
Robert Shoeman ◽  
...  
Keyword(s):  
Data Collection ◽  
Structure Factor ◽  
Free Electron ◽  
De Novo ◽  
Protein Crystallography ◽  
Pump Probe ◽  
Sample Data ◽  
Anomalous Signal ◽  
Beam Parameters

Free-electron lasers (FELs) are pushing back the limits of possibility in protein crystallography. Using the high-intensity, femtosecond duration pulses afforded by FELs allow data collection from micrometer-sized crystals while outrunning radiation damage. Moreover, FELs may be used for pump-probe experiments with unprecedented time resolution. However, the intricacies of FEL data collection pose specific challenges: as every FEL pulse destroys the sample, data are mostly collected from a stream of microcrystals and averaged to remove the variations in crystal size and quality as well as shot-to-shot variations in beam parameters. This technique is called serial femtosecond crystallography (SFX). In SFX, several tens of thousands of images typically need to be averaged to obtain reasonably accurate structure factor amplitudes. We previously showed that SFX yields structure factor amplitudes accurate enough to detect the weak anomalous signal of endogenous sulfur atoms. Now we show that SFX can be used to collect data accurate enough for de-novo phasing of a protein structure[1]. Using a model system (gadolinium-derivatized lysozyme) we collected ~60,000 diffraction images and obtained structure factor amplitudes that allowed phasing by single-wavelength anomalous diffraction. This first demonstration of de novo phasing from FEL data leads us to anticipate that FEL-based crystallography will become an important tool for the structure determination of proteins that are extremely radiation sensitive or that are difficult to crystallize, such as membrane proteins.

scholarly journals Protein structure determination by single-wavelength anomalous diffraction phasing of X-ray free-electron laser data

IUCrJ ◽  
2016 ◽  
Vol 3 (3) ◽  
pp. 180-191 ◽  
Author(s):  
Karol Nass ◽  
Anton Meinhart ◽  
Thomas R. M. Barends ◽  
Lutz Foucar ◽  
Alexander Gorel ◽  
...  
Keyword(s):  
Structure Determination ◽  
Free Electron ◽  
De Novo ◽  
Protein Structure Determination ◽  
Anomalous Scattering ◽  
X Ray ◽  
Sad Phasing ◽  
Anomalous Signal ◽  
Serial Femtosecond Crystallography

Serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs) offers unprecedented possibilities for macromolecular structure determination of systems that are prone to radiation damage. However, phasing XFEL datade novois complicated by the inherent inaccuracy of SFX data, and only a few successful examples, mostly based on exceedingly strong anomalous or isomorphous difference signals, have been reported. Here, it is shown that SFX data from thaumatin microcrystals can be successfully phased using only the weak anomalous scattering from the endogenous S atoms. Moreover, a step-by-step investigation is presented of the particular problems of SAD phasing of SFX data, analysing data from a derivative with a strong anomalous signal as well as the weak signal from endogenous S atoms.

scholarly journals ANALISIS KEBIJAKAN PENDIRIAN SEKOLAH DASAR DI KABUPATEN BATU BARA

2021 ◽  
Vol 11 (2) ◽  
pp. 102-107
Author(s):  
Regina Sipayung ◽  
◽  
Din Oloan Sihotang ◽  
Johannes Sohirimon Lumban Batu ◽  
◽  
...  
Keyword(s):  
Data Collection ◽  
Structure Factor ◽  
Interview Data ◽  
Communication Process ◽  
Study Group ◽  
National Education ◽  
School Conditions ◽  
Bureaucratic Structure

This study uses a descriptive methodology with interview data collection techniques, observation and documentation studies. The results showed that of the 17 schools that were established and categorized since 2018 it was found that 37% of the schools did not meet the study group requirements because there were less than 20 students. Then, judging from the communication process, the socialization of the requirements for the establishment of schools has not been carried out properly, information regarding the determination of school conditions is only conveyed to the school establishment committee. The readiness of resources has not been met and the disposition process still needs attention. The bureaucratic structure factor is still not clear, the standard operational procedures of the Batu Bara District Education Office have been guided by Kepmendiknas No. 060/U/2002. Based on the conclusion of this study, it was found that the Education Office of Batu Bara Regency had implemented policies in accordance with the Ministry of National Education. Number 060/U/2002, but needs to do some repairs. Keywords: Policy, School Establishment, Edwards

scholarly journals Enabling X-ray free electron laser crystallography for challenging biological systems from a limited number of crystals

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Monarin Uervirojnangkoorn ◽  
Oliver B Zeldin ◽  
Artem Y Lyubimov ◽  
Johan Hattne ◽  
Aaron S Brewster ◽  
...  
Keyword(s):  
Data Processing ◽  
Diffraction Data ◽  
Free Electron ◽  
Biological Systems ◽  
Processing System ◽  
Data Set ◽  
X Ray ◽  
Density Maps ◽  
Beam Parameters

There is considerable potential for X-ray free electron lasers (XFELs) to enable determination of macromolecular crystal structures that are difficult to solve using current synchrotron sources. Prior XFEL studies often involved the collection of thousands to millions of diffraction images, in part due to limitations of data processing methods. We implemented a data processing system based on classical post-refinement techniques, adapted to specific properties of XFEL diffraction data. When applied to XFEL data from three different proteins collected using various sample delivery systems and XFEL beam parameters, our method improved the quality of the diffraction data as well as the resulting refined atomic models and electron density maps. Moreover, the number of observations for a reflection necessary to assemble an accurate data set could be reduced to a few observations. These developments will help expand the applicability of XFEL crystallography to challenging biological systems, including cases where sample is limited.

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 Native phasing of x-ray free-electron laser data for a G protein–coupled receptor

2016 ◽  
Vol 2 (9) ◽  
pp. e1600292 ◽  
Author(s):  
Alexander Batyuk ◽  
Lorenzo Galli ◽  
Andrii Ishchenko ◽  
Gye Won Han ◽  
Cornelius Gati ◽  
...  
Keyword(s):  
Free Electron ◽  
Room Temperature ◽  
De Novo ◽  
Ultrashort Pulses ◽  
Temperature Structure ◽  
X Ray ◽  
Anomalous Signal ◽  
G Protein Coupled ◽  
Serial Femtosecond Crystallography ◽  
General Method

Serial femtosecond crystallography (SFX) takes advantage of extremely bright and ultrashort pulses produced by x-ray free-electron lasers (XFELs), allowing for the collection of high-resolution diffraction intensities from micrometer-sized crystals at room temperature with minimal radiation damage, using the principle of “diffraction-before-destruction.” However, de novo structure factor phase determination using XFELs has been difficult so far. We demonstrate the ability to solve the crystallographic phase problem for SFX data collected with an XFEL using the anomalous signal from native sulfur atoms, leading to a bias-free room temperature structure of the human A2A adenosine receptor at 1.9 Å resolution. The advancement was made possible by recent improvements in SFX data analysis and the design of injectors and delivery media for streaming hydrated microcrystals. This general method should accelerate structural studies of novel difficult-to-crystallize macromolecules and their complexes.

scholarly journals Experimental phase determination with selenomethionine or mercury-derivatization in serial femtosecond crystallography

IUCrJ ◽  
2017 ◽  
Vol 4 (5) ◽  
pp. 639-647 ◽  
Author(s):  
Keitaro Yamashita ◽  
Naoyuki Kuwabara ◽  
Takanori Nakane ◽  
Tomohiro Murai ◽  
Eiichi Mizohata ◽  
...  
Keyword(s):  
Structure Determination ◽  
Free Electron ◽  
De Novo ◽  
Heavy Atom ◽  
Protein Structure Determination ◽  
High Energy ◽  
X Rays ◽  
Replacement Method ◽  
Serial Femtosecond Crystallography

Serial femtosecond crystallography (SFX) using X-ray free-electron lasers (XFELs) holds enormous potential for the structure determination of proteins for which it is difficult to produce large and high-quality crystals. SFX has been applied to various systems, but rarely to proteins that have previously unknown structures. Consequently, the majority of previously obtained SFX structures have been solved by the molecular replacement method. To facilitate protein structure determination by SFX, it is essential to establish phasing methods that work efficiently for SFX. Here, selenomethionine derivatization and mercury soaking have been investigated for SFX experiments using the high-energy XFEL at the SPring-8 Angstrom Compact Free-Electron Laser (SACLA), Hyogo, Japan. Three successful cases are reported of single-wavelength anomalous diffraction (SAD) phasing using X-rays of less than 1 Å wavelength with reasonable numbers of diffraction patterns (13 000, 60 000 and 11 000). It is demonstrated that the combination of high-energy X-rays from an XFEL and commonly used heavy-atom incorporation techniques will enable routinede novostructural determination of biomacromolecules.

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.

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.

scholarly journals Effective data collection and refinement of pertubated structures

2014 ◽  
Vol 70 (a1) ◽  
pp. C1750-C1750
Author(s):  
Pascal Parois ◽  
Richard Cooper
Keyword(s):  
Excited State ◽  
Data Collection ◽  
Least Squares ◽  
Time Resolved ◽  
Pump Probe ◽  
Partial Data ◽  
Mixed Oxidation ◽  
Probe Experiment ◽  
Pump Probe Experiment

Approaches to determining the influence of individual measurements on the precision of crystallographic least squares parameters have been known for a long while.[1] Situations in which the precision of a single parameter (or linear combination of parameters) is critical can include: determination of novel bond lengths; refinement of site occupancies in mixed metal or mixed oxidation state systems; determination of the fraction of excited state molecules in a time-resolved pump-probe experiment. Such calculations are easily applicable to point-detector instruments, where individual influential reflections could be remeasured one-by-one. However, on a modern area detector instrument many reflections are measured on one frame and therefore some consideration of the appropriate strategy of reciprocal space scans is permitted to allow a more efficient use of the instrument. The highly influential partial data collection is then feed into an appropriate refinement model. Occupancies in mixed-metal or mixed-oxidation state systems and fractions and positions of excited state molecules during a time-resolved pump-probe experiment can be determined using direct refinement of the perturbation of the structure from the ground state. Re-factoring to modern Fortran of the Crystals software is in progress to allow the implementation of new algorithms such as a difference refinement.[2] We present an analysis of diffractometer strategy selection to prioritize scans which give the best improvement in specific least-squares parameters and a novel algorithm for the refinement of the partial data using crystals.

ANALISIS NILAI KOHESI DAN KOHERENSIDALAM TERJEMAHAN AL-QUR’AN SURAH AL AL ZALZALAH

2016 ◽  
Vol 1 (2) ◽  
pp. 23
Author(s):  
MUNIRAH MUNIRAH ◽  
HUSAIN SYARIFUDDIN
Keyword(s):  
Data Analysis ◽  
Data Collection ◽  
Data Classification ◽  
Research Data ◽  
Relative Pronoun ◽  
Second Person ◽  
Record Keeping ◽  
Descriptive Research ◽  
Qualitative Descriptive

This study aimed to describe the value of cohesion and coherence contained in the translation of the Qur'an surah Al Zalzalah. This study was a qualitative descriptive research, research data collection techniques using three techniques namely, inventory, rading and understanding, and record keeping. The data analysis used the coding of data, classification data, and the determination of the data. The results showed that the cohesion markers used in the translation of surah Al Zalzalah discourse are: 1) reference, 2) pronouns, ie pronouns second person, and third, the relative pronoun, the pronoun pointer, pen pronouns and pronouns owner, 3 ) conjunctions, namely temporal conjunctions, coordinating conjunctions, subordinating conjunctions, and conjunctions koorelatif, and 4) a causal ellipsis. It mean that there was a coherence in the translation of surah Al Zalzalah discourse are: the addition or addition, pronouns, repetition or repetition, match words or synonyms, in whole or in part, a comparison or ratio of conclusions or results. Keywords: Cohesion, Coherence, sura Al Zalzalah AbstrakPenelitian ini bertujuan untuk mendeskripsikan nilai kohesi dan koherensi yang terdapat dalam terjemahan Al-Qur’an surah Al Zalzalah. Jenis penelitian ini termasuk jenis penelitian deskriptif kualitatif, Teknik pengumpulan data penelitian menggunakan tiga teknik yakni, inventarisasi, baca simak, dan pencatatan. Teknik analisis data menggunakan pengodean data, pengklasifikasian data, dan penentuan data. Hasil penelitian menunjukkan bahwa pemarkah kohesi yang digunakan dalam wacana terjemahan surah Al Zalzalah adalah: 1) referensi, 2) pronomina, yaitu kata ganti orang kedua, dan ketiga, kata ganti penghubung, kata ganti penunjuk, kata ganti penanya dan kata ganti empunya, 3) konjungsi, yaitu konjungsi temporal, konjungsi koordinatif, konjungsi subordinatif, dan konjungsi koorelatif, dan 4) elipsis kausal. Sarana koherensi yang terdapat di dalam wacana terjemahan surah Al Zalzalah adalah: penambahan atau adisi, pronomina, pengulangan atau repetisi, padan kata atau sinonim, keseluruhan atau bagian, komparasi atau perbandingan simpulan atau hasil.Kata Kunci: Kohesi, Koherensi, surah Al Zalzalah

Sign in / Sign up

Export Citation Format

Share Document