Publications by authors named "Johan Bielecki"

27 Publications

  • Page 1 of 1

Diffraction data from aerosolized Coliphage PR772 virus particles imaged with the Linac Coherent Light Source.

Sci Data 2020 11 19;7(1):404. Epub 2020 Nov 19.

Arizona State University, 1001S. McAllister Avenue, Tempe, AZ, 85287, USA.

Single Particle Imaging (SPI) with intense coherent X-ray pulses from X-ray free-electron lasers (XFELs) has the potential to produce molecular structures without the need for crystallization or freezing. Here we present a dataset of 285,944 diffraction patterns from aerosolized Coliphage PR772 virus particles injected into the femtosecond X-ray pulses of the Linac Coherent Light Source (LCLS). Additional exposures with background information are also deposited. The diffraction data were collected at the Atomic, Molecular and Optical Science Instrument (AMO) of the LCLS in 4 experimental beam times during a period of four years. The photon energy was either 1.2 or 1.7 keV and the pulse energy was between 2 and 4 mJ in a focal spot of about 1.3 μm x 1.7 μm full width at half maximum (FWHM). The X-ray laser pulses captured the particles in random orientations. The data offer insight into aerosolised virus particles in the gas phase, contain information relevant to improving experimental parameters, and provide a basis for developing algorithms for image analysis and reconstruction.
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http://dx.doi.org/10.1038/s41597-020-00745-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7678860PMC
November 2020

Segmented flow generator for serial crystallography at the European X-ray free electron laser.

Nat Commun 2020 09 9;11(1):4511. Epub 2020 Sep 9.

School of Molecular Sciences, Arizona State University, Tempe, AZ, 85287-1604, USA.

Serial femtosecond crystallography (SFX) with X-ray free electron lasers (XFELs) allows structure determination of membrane proteins and time-resolved crystallography. Common liquid sample delivery continuously jets the protein crystal suspension into the path of the XFEL, wasting a vast amount of sample due to the pulsed nature of all current XFEL sources. The European XFEL (EuXFEL) delivers femtosecond (fs) X-ray pulses in trains spaced 100 ms apart whereas pulses within trains are currently separated by 889 ns. Therefore, continuous sample delivery via fast jets wastes >99% of sample. Here, we introduce a microfluidic device delivering crystal laden droplets segmented with an immiscible oil reducing sample waste and demonstrate droplet injection at the EuXFEL compatible with high pressure liquid delivery of an SFX experiment. While achieving ~60% reduction in sample waste, we determine the structure of the enzyme 3-deoxy-D-manno-octulosonate-8-phosphate synthase from microcrystals delivered in droplets revealing distinct structural features not previously reported.
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http://dx.doi.org/10.1038/s41467-020-18156-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7481229PMC
September 2020

Perspectives on single particle imaging with x rays at the advent of high repetition rate x-ray free electron laser sources.

Struct Dyn 2020 Jul 6;7(4):040901. Epub 2020 Aug 6.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), SE-75124 Uppsala, Sweden.

X-ray free electron lasers (XFELs) now routinely produce millijoule level pulses of x-ray photons with tens of femtoseconds duration. Such x-ray intensities gave rise to the idea that weakly scattering particles-perhaps single biomolecules or viruses-could be investigated free of radiation damage. Here, we examine elements from the past decade of so-called single particle imaging with hard XFELs. We look at the progress made to date and identify some future possible directions for the field. In particular, we summarize the presently achieved resolutions as well as identifying the bottlenecks and enabling technologies to future resolution improvement, which in turn enables application to samples of scientific interest.
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http://dx.doi.org/10.1063/4.0000024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7413746PMC
July 2020

Author Correction: Membrane protein megahertz crystallography at the European XFEL.

Nat Commun 2020 Jan 30;11(1):703. Epub 2020 Jan 30.

Biodesign Center for Applied Structural Discovery, Arizona State University, Tempe, AZ, 85287-5001, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41467-020-14436-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6992783PMC
January 2020

The role of transient resonances for ultra-fast imaging of single sucrose nanoclusters.

Nat Commun 2020 01 9;11(1):167. Epub 2020 Jan 9.

Chemical Sciences and Engineering Division, Argonne National Laboratory, Argonne, IL, 60439, USA.

Intense x-ray free-electron laser (XFEL) pulses hold great promise for imaging function in nanoscale and biological systems with atomic resolution. So far, however, the spatial resolution obtained from single shot experiments lags averaging static experiments. Here we report on a combined computational and experimental study about ultrafast diffractive imaging of sucrose clusters which are benchmark organic samples. Our theoretical model matches the experimental data from the water window to the keV x-ray regime. The large-scale dynamic scattering calculations reveal that transient phenomena driven by non-linear x-ray interaction are decisive for ultrafast imaging applications. Our study illuminates the complex interplay of the imaging process with the rapidly changing transient electronic structures in XFEL experiments and shows how computational models allow optimization of the parameters for ultrafast imaging experiments.
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http://dx.doi.org/10.1038/s41467-019-13905-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952381PMC
January 2020

Time-resolved serial femtosecond crystallography at the European XFEL.

Nat Methods 2020 01 18;17(1):73-78. Epub 2019 Nov 18.

Physics Department, University of Wisconsin-Milwaukee, Milwaukee, WI, USA.

The European XFEL (EuXFEL) is a 3.4-km long X-ray source, which produces femtosecond, ultrabrilliant and spatially coherent X-ray pulses at megahertz (MHz) repetition rates. This X-ray source has been designed to enable the observation of ultrafast processes with near-atomic spatial resolution. Time-resolved crystallographic investigations on biological macromolecules belong to an important class of experiments that explore fundamental and functional structural displacements in these molecules. Due to the unusual MHz X-ray pulse structure at the EuXFEL, these experiments are challenging. Here, we demonstrate how a biological reaction can be followed on ultrafast timescales at the EuXFEL. We investigate the picosecond time range in the photocycle of photoactive yellow protein (PYP) with MHz X-ray pulse rates. We show that difference electron density maps of excellent quality can be obtained. The results connect the previously explored femtosecond PYP dynamics to timescales accessible at synchrotrons. This opens the door to a wide range of time-resolved studies at the EuXFEL.
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http://dx.doi.org/10.1038/s41592-019-0628-zDOI Listing
January 2020

Membrane protein megahertz crystallography at the European XFEL.

Nat Commun 2019 11 4;10(1):5021. Epub 2019 Nov 4.

Biodesign Center for Applied Structural Discovery, Arizona State University, Tempe, AZ, 85287-5001, USA.

The world's first superconducting megahertz repetition rate hard X-ray free-electron laser (XFEL), the European XFEL, began operation in 2017, featuring a unique pulse train structure with 886 ns between pulses. With its rapid pulse rate, the European XFEL may alleviate some of the increasing demand for XFEL beamtime, particularly for membrane protein serial femtosecond crystallography (SFX), leveraging orders-of-magnitude faster data collection. Here, we report the first membrane protein megahertz SFX experiment, where we determined a 2.9 Å-resolution SFX structure of the large membrane protein complex, Photosystem I, a > 1 MDa complex containing 36 protein subunits and 381 cofactors. We address challenges to megahertz SFX for membrane protein complexes, including growth of large quantities of crystals and the large molecular and unit cell size that influence data collection and analysis. The results imply that megahertz crystallography could have an important impact on structure determination of large protein complexes with XFELs.
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http://dx.doi.org/10.1038/s41467-019-12955-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6828683PMC
November 2019

The Single Particles, Clusters and Biomolecules and Serial Femtosecond Crystallography instrument of the European XFEL: initial installation.

J Synchrotron Radiat 2019 May 12;26(Pt 3):660-676. Epub 2019 Apr 12.

European XFEL, Holzkoppel 4, 22869 Schenefeld, Germany.

The European X-ray Free-Electron Laser (FEL) became the first operational high-repetition-rate hard X-ray FEL with first lasing in May 2017. Biological structure determination has already benefitted from the unique properties and capabilities of X-ray FELs, predominantly through the development and application of serial crystallography. The possibility of now performing such experiments at data rates more than an order of magnitude greater than previous X-ray FELs enables not only a higher rate of discovery but also new classes of experiments previously not feasible at lower data rates. One example is time-resolved experiments requiring a higher number of time steps for interpretation, or structure determination from samples with low hit rates in conventional X-ray FEL serial crystallography. Following first lasing at the European XFEL, initial commissioning and operation occurred at two scientific instruments, one of which is the Single Particles, Clusters and Biomolecules and Serial Femtosecond Crystallography (SPB/SFX) instrument. This instrument provides a photon energy range, focal spot sizes and diagnostic tools necessary for structure determination of biological specimens. The instrumentation explicitly addresses serial crystallography and the developing single particle imaging method as well as other forward-scattering and diffraction techniques. This paper describes the major science cases of SPB/SFX and its initial instrumentation - in particular its optical systems, available sample delivery methods, 2D detectors, supporting optical laser systems and key diagnostic components. The present capabilities of the instrument will be reviewed and a brief outlook of its future capabilities is also described.
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http://dx.doi.org/10.1107/S1600577519003308DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6510195PMC
May 2019

Electrospray sample injection for single-particle imaging with x-ray lasers.

Sci Adv 2019 05 3;5(5):eaav8801. Epub 2019 May 3.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), SE-75124 Uppsala, Sweden.

The possibility of imaging single proteins constitutes an exciting challenge for x-ray lasers. Despite encouraging results on large particles, imaging small particles has proven to be difficult for two reasons: not quite high enough pulse intensity from currently available x-ray lasers and, as we demonstrate here, contamination of the aerosolized molecules by nonvolatile contaminants in the solution. The amount of contamination on the sample depends on the initial droplet size during aerosolization. Here, we show that, with our electrospray injector, we can decrease the size of aerosol droplets and demonstrate virtually contaminant-free sample delivery of organelles, small virions, and proteins. The results presented here, together with the increased performance of next-generation x-ray lasers, constitute an important stepping stone toward the ultimate goal of protein structure determination from imaging at room temperature and high temporal resolution.
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http://dx.doi.org/10.1126/sciadv.aav8801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6499549PMC
May 2019

MHz data collection of a microcrystalline mixture of different jack bean proteins.

Sci Data 2019 04 3;6(1):18. Epub 2019 Apr 3.

Max Planck Institute for Medical Research, Jahnstrasse 29, 69120, Heidelberg, Germany.

We provide a detailed description of a serial femtosecond crystallography (SFX) dataset collected at the European X-ray free-electron laser facility (EuXFEL). The EuXFEL is the first high repetition rate XFEL delivering MHz X-ray pulse trains at 10 Hz. The short spacing (<1 µs) between pulses requires fast flowing microjets for sample injection and high frame rate detectors. A data set was recorded of a microcrystalline mixture of at least three different jack bean proteins (urease, concanavalin A, concanavalin B). A one megapixel Adaptive Gain Integrating Pixel Detector (AGIPD) was used which has not only a high frame rate but also a large dynamic range. This dataset is publicly available through the Coherent X-ray Imaging Data Bank (CXIDB) as a resource for algorithm development and for data analysis training for prospective XFEL users.
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http://dx.doi.org/10.1038/s41597-019-0010-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472352PMC
April 2019

Single-particle imaging without symmetry constraints at an X-ray free-electron laser.

IUCrJ 2018 Nov 18;5(Pt 6):727-736. Epub 2018 Sep 18.

Deutsches Elektronen-Synchrotron DESY, Notkestrasse 85, Hamburg D-22607, Germany.

The analysis of a single-particle imaging (SPI) experiment performed at the AMO beamline at LCLS as part of the SPI initiative is presented here. A workflow for the three-dimensional virus reconstruction of the PR772 bacteriophage from measured single-particle data is developed. It consists of several well defined steps including single-hit diffraction data classification, refined filtering of the classified data, reconstruction of three-dimensional scattered intensity from the experimental diffraction patterns by orientation determination and a final three-dimensional reconstruction of the virus electron density without symmetry constraints. The analysis developed here revealed and quantified nanoscale features of the PR772 virus measured in this experiment, with the obtained resolution better than 10 nm, with a clear indication that the structure was compressed in one direction and, as such, deviates from ideal icosahedral symmetry.
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http://dx.doi.org/10.1107/S205225251801120XDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6211532PMC
November 2018

Rayleigh-scattering microscopy for tracking and sizing nanoparticles in focused aerosol beams.

IUCrJ 2018 Nov 11;5(Pt 6):673-680. Epub 2018 Sep 11.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), Uppsala SE-75124, Sweden.

Ultra-bright femtosecond X-ray pulses generated by X-ray free-electron lasers (XFELs) can be used to image high-resolution structures without the need for crystallization. For this approach, aerosol injection has been a successful method to deliver 70-2000 nm particles into the XFEL beam efficiently and at low noise. Improving the technique of aerosol sample delivery and extending it to single proteins necessitates quantitative aerosol diagnostics. Here a lab-based technique is introduced for Rayleigh-scattering microscopy allowing us to track and size aerosolized particles down to 40 nm in diameter as they exit the injector. This technique was used to characterize the 'Uppsala injector', which is a pioneering and frequently used aerosol sample injector for XFEL single-particle imaging. The particle-beam focus, particle velocities, particle density and injection yield were measured at different operating conditions. It is also shown how high particle densities and good injection yields can be reached for large particles (100-500 nm). It is found that with decreasing particle size, particle densities and injection yields deteriorate, indicating the need for different injection strategies to extend XFEL imaging to smaller targets, such as single proteins. This work demonstrates the power of Rayleigh-scattering microscopy for studying focused aerosol beams quantitatively. It lays the foundation for lab-based injector development and online injection diagnostics for XFEL research. In the future, the technique may also find application in other fields that employ focused aerosol beams, such as mass spectrometry, particle deposition, fuel injection and three-dimensional printing techniques.
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http://dx.doi.org/10.1107/S2052252518010837DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6211534PMC
November 2018

Megahertz serial crystallography.

Authors:
Max O Wiedorn Dominik Oberthür Richard Bean Robin Schubert Nadine Werner Brian Abbey Martin Aepfelbacher Luigi Adriano Aschkan Allahgholi Nasser Al-Qudami Jakob Andreasson Steve Aplin Salah Awel Kartik Ayyer Saša Bajt Imrich Barák Sadia Bari Johan Bielecki Sabine Botha Djelloul Boukhelef Wolfgang Brehm Sandor Brockhauser Igor Cheviakov Matthew A Coleman Francisco Cruz-Mazo Cyril Danilevski Connie Darmanin R Bruce Doak Martin Domaracky Katerina Dörner Yang Du Hans Fangohr Holger Fleckenstein Matthias Frank Petra Fromme Alfonso M Gañán-Calvo Yaroslav Gevorkov Klaus Giewekemeyer Helen Mary Ginn Heinz Graafsma Rita Graceffa Dominic Greiffenberg Lars Gumprecht Peter Göttlicher Janos Hajdu Steffen Hauf Michael Heymann Susannah Holmes Daniel A Horke Mark S Hunter Siegfried Imlau Alexander Kaukher Yoonhee Kim Alexander Klyuev Juraj Knoška Bostjan Kobe Manuela Kuhn Christopher Kupitz Jochen Küpper Janine Mia Lahey-Rudolph Torsten Laurus Karoline Le Cong Romain Letrun P Lourdu Xavier Luis Maia Filipe R N C Maia Valerio Mariani Marc Messerschmidt Markus Metz Davide Mezza Thomas Michelat Grant Mills Diana C F Monteiro Andrew Morgan Kerstin Mühlig Anna Munke Astrid Münnich Julia Nette Keith A Nugent Theresa Nuguid Allen M Orville Suraj Pandey Gisel Pena Pablo Villanueva-Perez Jennifer Poehlsen Gianpietro Previtali Lars Redecke Winnie Maria Riekehr Holger Rohde Adam Round Tatiana Safenreiter Iosifina Sarrou Tokushi Sato Marius Schmidt Bernd Schmitt Robert Schönherr Joachim Schulz Jonas A Sellberg M Marvin Seibert Carolin Seuring Megan L Shelby Robert L Shoeman Marcin Sikorski Alessandro Silenzi Claudiu A Stan Xintian Shi Stephan Stern Jola Sztuk-Dambietz Janusz Szuba Aleksandra Tolstikova Martin Trebbin Ulrich Trunk Patrik Vagovic Thomas Ve Britta Weinhausen Thomas A White Krzysztof Wrona Chen Xu Oleksandr Yefanov Nadia Zatsepin Jiaguo Zhang Markus Perbandt Adrian P Mancuso Christian Betzel Henry Chapman Anton Barty

Nat Commun 2018 10 2;9(1):4025. Epub 2018 Oct 2.

Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, Notkestrasse 85, 22607, Hamburg, Germany.

The new European X-ray Free-Electron Laser is the first X-ray free-electron laser capable of delivering X-ray pulses with a megahertz inter-pulse spacing, more than four orders of magnitude higher than previously possible. However, to date, it has been unclear whether it would indeed be possible to measure high-quality diffraction data at megahertz pulse repetition rates. Here, we show that high-quality structures can indeed be obtained using currently available operating conditions at the European XFEL. We present two complete data sets, one from the well-known model system lysozyme and the other from a so far unknown complex of a β-lactamase from K. pneumoniae involved in antibiotic resistance. This result opens up megahertz serial femtosecond crystallography (SFX) as a tool for reliable structure determination, substrate screening and the efficient measurement of the evolution and dynamics of molecular structures using megahertz repetition rate pulses available at this new class of X-ray laser source.
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http://dx.doi.org/10.1038/s41467-018-06156-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6168542PMC
October 2018

Considerations for three-dimensional image reconstruction from experimental data in coherent diffractive imaging.

IUCrJ 2018 Sep 1;5(Pt 5):531-541. Epub 2018 Sep 1.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), SE-751 24 Uppsala, Sweden.

Diffraction before destruction using X-ray free-electron lasers (XFELs) has the potential to determine radiation-damage-free structures without the need for crystallization. This article presents the three-dimensional reconstruction of the Melbournevirus from single-particle X-ray diffraction patterns collected at the LINAC Coherent Light Source (LCLS) as well as reconstructions from simulated data exploring the consequences of different kinds of experimental sources of noise. The reconstruction from experimental data suffers from a strong artifact in the center of the particle. This could be reproduced with simulated data by adding experimental background to the diffraction patterns. In those simulations, the relative density of the artifact increases linearly with background strength. This suggests that the artifact originates from the Fourier transform of the relatively flat background, concentrating all power in a central feature of limited extent. We support these findings by significantly reducing the artifact through background removal before the phase-retrieval step. Large amounts of blurring in the diffraction patterns were also found to introduce diffuse artifacts, which could easily be mistaken as biologically relevant features. Other sources of noise such as sample heterogeneity and variation of pulse energy did not significantly degrade the quality of the reconstructions. Larger data volumes, made possible by the recent inauguration of high repetition-rate XFELs, allow for increased signal-to-background ratio and provide a way to minimize these artifacts. The anticipated development of three-dimensional Fourier-volume-assembly algorithms which are background aware is an alternative and complementary solution, which maximizes the use of data.
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http://dx.doi.org/10.1107/S2052252518010047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6126651PMC
September 2018

Megahertz data collection from protein microcrystals at an X-ray free-electron laser.

Nat Commun 2018 08 28;9(1):3487. Epub 2018 Aug 28.

Max Planck Institute for Medical Research, Jahnstrasse 29, 69120, Heidelberg, Germany.

X-ray free-electron lasers (XFELs) enable novel experiments because of their high peak brilliance and femtosecond pulse duration. However, non-superconducting XFELs offer repetition rates of only 10-120 Hz, placing significant demands on beam time and sample consumption. We describe serial femtosecond crystallography experiments performed at the European XFEL, the first MHz repetition rate XFEL, delivering 1.128 MHz X-ray pulse trains at 10 Hz. Given the short spacing between pulses, damage caused by shock waves launched by one XFEL pulse on sample probed by subsequent pulses is a concern. To investigate this issue, we collected data from lysozyme microcrystals, exposed to a ~15 μm XFEL beam. Under these conditions, data quality is independent of whether the first or subsequent pulses of the train were used for data collection. We also analyzed a mixture of microcrystals of jack bean proteins, from which the structure of native, magnesium-containing concanavalin A was determined.
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http://dx.doi.org/10.1038/s41467-018-05953-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6113309PMC
August 2018

Correlations in Scattered X-Ray Laser Pulses Reveal Nanoscale Structural Features of Viruses.

Phys Rev Lett 2017 Oct 12;119(15):158102. Epub 2017 Oct 12.

European XFEL GmbH, Holzkoppel 4, D-22869 Schenefeld, Germany.

We use extremely bright and ultrashort pulses from an x-ray free-electron laser (XFEL) to measure correlations in x rays scattered from individual bioparticles. This allows us to go beyond the traditional crystallography and single-particle imaging approaches for structure investigations. We employ angular correlations to recover the three-dimensional (3D) structure of nanoscale viruses from x-ray diffraction data measured at the Linac Coherent Light Source. Correlations provide us with a comprehensive structural fingerprint of a 3D virus, which we use both for model-based and ab initio structure recovery. The analyses reveal a clear indication that the structure of the viruses deviates from the expected perfect icosahedral symmetry. Our results anticipate exciting opportunities for XFEL studies of the structure and dynamics of nanoscale objects by means of angular correlations.
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http://dx.doi.org/10.1103/PhysRevLett.119.158102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5757528PMC
October 2017

Coherent soft X-ray diffraction imaging of coliphage PR772 at the Linac coherent light source.

Sci Data 2017 06 27;4:170079. Epub 2017 Jun 27.

Center for Free-Electron Laser Science, DESY, Notkestrasse 85, 22607 Hamburg, Germany.

Single-particle diffraction from X-ray Free Electron Lasers offers the potential for molecular structure determination without the need for crystallization. In an effort to further develop the technique, we present a dataset of coherent soft X-ray diffraction images of Coliphage PR772 virus, collected at the Atomic Molecular Optics (AMO) beamline with pnCCD detectors in the LAMP instrument at the Linac Coherent Light Source. The diameter of PR772 ranges from 65-70 nm, which is considerably smaller than the previously reported ~600 nm diameter Mimivirus. This reflects continued progress in XFEL-based single-particle imaging towards the single molecular imaging regime. The data set contains significantly more single particle hits than collected in previous experiments, enabling the development of improved statistical analysis, reconstruction algorithms, and quantitative metrics to determine resolution and self-consistency.
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http://dx.doi.org/10.1038/sdata.2017.79DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5501160PMC
June 2017

Experimental strategies for imaging bioparticles with femtosecond hard X-ray pulses.

IUCrJ 2017 May 7;4(Pt 3):251-262. Epub 2017 Apr 7.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), SE-751 24 Uppsala, Sweden.

This study explores the capabilities of the Coherent X-ray Imaging Instrument at the Linac Coherent Light Source to image small biological samples. The weak signal from small samples puts a significant demand on the experiment. Aerosolized particles of ∼40 nm in diameter were injected into the submicrometre X-ray focus at a reduced pressure. Diffraction patterns were recorded on two area detectors. The statistical nature of the measurements from many individual particles provided information about the intensity profile of the X-ray beam, phase variations in the wavefront and the size distribution of the injected particles. The results point to a wider than expected size distribution (from ∼35 to ∼300 nm in diameter). This is likely to be owing to nonvolatile contaminants from larger droplets during aerosolization and droplet evaporation. The results suggest that the concentration of nonvolatile contaminants and the ratio between the volumes of the initial droplet and the sample particles is critical in such studies. The maximum beam intensity in the focus was found to be 1.9 × 10 photons per µm per pulse. The full-width of the focus at half-maximum was estimated to be 500 nm (assuming 20% beamline transmission), and this width is larger than expected. Under these conditions, the diffraction signal from a sample-sized particle remained above the average background to a resolution of 4.25 nm. The results suggest that reducing the size of the initial droplets during aerosolization is necessary to bring small particles into the scope of detailed structural studies with X-ray lasers.
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http://dx.doi.org/10.1107/S2052252517003591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5414399PMC
May 2017

A data set from flash X-ray imaging of carboxysomes.

Sci Data 2016 Aug 1;3:160061. Epub 2016 Aug 1.

Department of Cell and Molecular Biology, Laboratory of Molecular Biophysics, Uppsala University, Husargatan 3 (Box 596), Uppsala SE-751 24, Sweden.

Ultra-intense femtosecond X-ray pulses from X-ray lasers permit structural studies on single particles and biomolecules without crystals. We present a large data set on inherently heterogeneous, polyhedral carboxysome particles. Carboxysomes are cell organelles that vary in size and facilitate up to 40% of Earth's carbon fixation by cyanobacteria and certain proteobacteria. Variation in size hinders crystallization. Carboxysomes appear icosahedral in the electron microscope. A protein shell encapsulates a large number of Rubisco molecules in paracrystalline arrays inside the organelle. We used carboxysomes with a mean diameter of 115±26 nm from Halothiobacillus neapolitanus. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min. Every diffraction pattern is a unique structure measurement and high-throughput imaging allows sampling the space of structural variability. The different structures can be separated and phased directly from the diffraction data and open a way for accurate, high-throughput studies on structures and structural heterogeneity in biology and elsewhere.
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http://dx.doi.org/10.1038/sdata.2016.61DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4968195PMC
August 2016

Open data set of live cyanobacterial cells imaged using an X-ray laser.

Sci Data 2016 Aug 1;3:160058. Epub 2016 Aug 1.

Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, Husargatan 3 (Box 596), SE-751 24 Uppsala, Sweden.

Structural studies on living cells by conventional methods are limited to low resolution because radiation damage kills cells long before the necessary dose for high resolution can be delivered. X-ray free-electron lasers circumvent this problem by outrunning key damage processes with an ultra-short and extremely bright coherent X-ray pulse. Diffraction-before-destruction experiments provide high-resolution data from cells that are alive when the femtosecond X-ray pulse traverses the sample. This paper presents two data sets from micron-sized cyanobacteria obtained at the Linac Coherent Light Source, containing a total of 199,000 diffraction patterns. Utilizing this type of diffraction data will require the development of new analysis methods and algorithms for studying structure and structural variability in large populations of cells and to create abstract models. Such studies will allow us to understand living cells and populations of cells in new ways. New X-ray lasers, like the European XFEL, will produce billions of pulses per day, and could open new areas in structural sciences.
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http://dx.doi.org/10.1038/sdata.2016.58DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4968219PMC
August 2016

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

Sci Data 2016 08 1;3:160064. Epub 2016 Aug 1.

Arizona State University, School of Molecular Sciences (SMS), Tempe, Arizona 85287-1604, USA.

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 well-characterized 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 μm diameter focused hard X-ray beam at the CXI instrument of LCLS. Diffraction patterns from RDV with signal to 5.9 Ångström 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.
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http://dx.doi.org/10.1038/sdata.2016.64DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4968191PMC
August 2016

The effect of alendronate on biomineralization at the bone/implant interface.

J Biomed Mater Res A 2016 03 12;104(3):620-629. Epub 2015 Nov 12.

Department of Chemistry and Chemical Engineering, Chalmers University of Technology, Gothenburg, Sweden.

A recent approach to improve the osseointegration of implants is to utilize local drug administration. The presence of an osteoporosis drug may influence both bone quantity and quality at the bone/implant interface. Despite this, the performance of bone-anchoring implants is traditionally evaluated only by quantitative measurements. In the present study, the osteoporosis drug alendronate (ALN) was administrated from mesoporous titania thin films that were coated onto titanium implants. The effect that the drug had on biomineralization was explored both in vitro using simulated body fluid (SBF) and in vivo in a rat tibia model. The SBF study showed that the apatite formation was completely hindered at a high concentration of ALN (0.1 mg/mL). However, when ALN was administrated from the mesoporous coating the surface became completely covered with apatite. Ex vivo characterization of the bone/implant interface using Raman spectroscopy demonstrated that the presence of ALN enhanced the bone mineralization, and that the chemical signature of newly formed bone in the presence of ALN had a higher resemblance to the pre-existing mature bone than to the bone formed without drug. Taken together, this study demonstrates the importance of evaluating the quality of the formed bone to better understand the performance of implants. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A 104A: 620-629, 2016.
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http://dx.doi.org/10.1002/jbm.a.35602DOI Listing
March 2016

Explosion dynamics of sucrose nanospheres monitored by time of flight spectrometry and coherent diffractive imaging at the split-and-delay beam line of the FLASH soft X-ray laser.

Opt Express 2014 Nov;22(23):28914-25

We use a Mach-Zehnder type autocorrelator to split and delay XUV pulses from the FLASH soft X-ray laser for triggering and subsequently probing the explosion of aerosolised sugar balls. FLASH was running at 182 eV photon energy with pulses of 70 fs duration. The delay between the pump-probe pulses was varied between zero and 5 ps, and the pulses were focused to reach peak intensities above 10¹⁶W/cm² with an off-axis parabola. The direct pulse triggered the explosion of single aerosolised sucrose nano-particles, while the delayed pulse probed the exploding structure. The ejected ions were measured by ion time of flight spectrometry, and the particle sizes were measured by coherent diffractive imaging. The results show that sucrose particles of 560-1000 nm diameter retain their size for about 500 fs following the first exposure. Significant sample expansion happens between 500 fs and 1 ps. We present simulations to support these observations.
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http://dx.doi.org/10.1364/OE.22.028914DOI Listing
November 2014

Automated identification and classification of single particle serial femtosecond X-ray diffraction data.

Opt Express 2014 Feb;22(3):2497-510

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.
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http://dx.doi.org/10.1364/OE.22.002497DOI Listing
February 2014

A statistical model of hydrogen bond networks in liquid alcohols.

J Chem Phys 2012 Mar;136(9):094514

Applied Physics, Chalmers University of Technology, SE-412 96 Göteborg, Sweden.

We here present a statistical model of hydrogen bond induced network structures in liquid alcohols. The model generalises the Andersson-Schulz-Flory chain model to allow also for branched structures. Two bonding probabilities are assigned to each hydroxyl group oxygen, where the first is the probability of a lone pair accepting an H-bond and the second is the probability that given this bond also the second lone pair is bonded. The average hydroxyl group cluster size, cluster size distribution, and the number of branches and leaves in the tree-like network clusters are directly determined from these probabilities. The applicability of the model is tested by comparison to cluster size distributions and bonding probabilities obtained from Monte Carlo simulations of the monoalcohols methanol, propanol, butanol, and propylene glycol monomethyl ether, the di-alcohol propylene glycol, and the tri-alcohol glycerol. We find that the tree model can reproduce the cluster size distributions and the bonding probabilities for both mono- and poly-alcohols, showing the branched nature of the OH-clusters in these liquids. Thus, this statistical model is a useful tool to better understand the structure of network forming hydrogen bonded liquids. The model can be applied to experimental data, allowing the topology of the clusters to be determined from such studies.
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http://dx.doi.org/10.1063/1.3690137DOI Listing
March 2012

Templated growth of covalently bonded three-dimensional carbon nanotube networks originated from graphene.

Adv Mater 2012 Mar 17;24(12):1576-81. Epub 2012 Feb 17.

Department of Microtechnology and Nanoscience, Chalmers University of Technology, Göteborg, Sweden.

A template-assisted method that enables the growth of covalently bonded three-dimensional carbon nanotubes (CNTs) originating from graphene at a large scale is demonstrated. Atomic force microscopy-based mechanical tests show that the covalently bonded CNT structure can effectively distribute external loading throughout the network to improve the mechanical strength of the material.
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http://dx.doi.org/10.1002/adma.201104408DOI Listing
March 2012