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Item type:Publication, Access status: with files , Iterative phase retrieval in macromolecular crystal diffraction using the gradients of intensities(International Union of Crystallography (IUCr), 2026-10-17); ; ; Iterative phase retrieval (IPR) offers a model-independent route to crystallographic structure determination but is fundamentally limited by the sampling of the diffraction pattern at Bragg positions. Consequently, conventional IPR methods typically rely on a high solvent fraction (>70%) to sufficiently constrain the solution, a condition met by only a small fraction of protein crystals. This work investigates a magnitude-and-gradient IPR algorithm that overcomes this undersampling barrier by exploiting the local intensity gradients ∇Iq observable in the diffraction patterns of finite crystals. By incorporating the gradients of the molecular transform alongside Bragg intensities, the phase problem is converted from an underdetermined system of equations to one that is overdetermined by a factor of two. Tests on simulated diffraction data for proteins in space group P1 demonstrate that the algorithm successfully reconstructs electron densities across a wide range of solvent fractions, including densely packed crystals where standard algorithms fail. Furthermore, the approach proves robust under experimentally relevant conditions, tolerating significant Poissonian noise and the loss of low-resolution terms due to a beamstop when coupled with approximate real-space support constraints. These results indicate that leveraging the fine structure of Bragg peaks at coherent X-ray sources can significantly extend the applicability of ab initio phasing.Publicationtype: Journal Article9 5 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Fast and efficient hard X-ray projection imaging below 10 nm resolution(Optica Publishing Group, 2024-08-12); ; ; High-resolution X-ray imaging of noncrystalline objects is often achieved through the approach of scanning coherent diffractive imaging known as ptychography. The imaging resolution is usually limited by the scattering properties of the sample, where weak diffraction signals at the highest scattering angles compete with parasitic scattering. Here, we demonstrate that X-ray multilayer Laue lenses with a high numerical aperture (NA) can be used to create a strong reference beam that holographically boosts weak scattering from the sample over a large range of scattering angles, enabling high-resolution imaging that is tolerant of such background. An imaging resolution of sub-10 nm was achieved at a photon energy of 17.4 keV with lenses of 0.014 NA from a Siemens star test object and a sample of hierarchical nanoporous gold, recording projection holograms at an effective magnification of more than 30,000 directly on a pixel-array detector. A numerical study compared this approach to low-NA far-field ptychography, indicating significant advantages for using high-NA lenses in the presence of background noise. This imaging modality is particularly fast and efficient at recording high-resolution transmission phase-contrast images over large fields of view in a facile manner.Publicationtype: Journal Article14 15 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , FDIP : a Fast Diffraction Image Processing library for X-ray crystallography experiments(Multidisciplinary Digital Publishing Institute (MDPI), 2024-02-05); ; ; ; Serial crystallography (SX) is a cutting-edge technique in structural biology, involving the systematic collection of X-ray diffraction data from numerous randomly oriented microcrystals. To extract comprehensive three-dimensional information about the studied system, SX utilises thousands of measured diffraction patterns. As such, SX takes advantages of the properties of modern X-ray sources, including Free Electron Lasers (FELs) and third and fourth generation synchrotrons, as well as contemporary high-repetition-rate detectors. Efficient analysis of the extensive datasets generated during SX experiments demands fast and effective algorithms. The FDIP library offers meticulously optimised functions tailored for preprocessing data obtained in SX experiments. This encompasses tasks such as background subtraction, identification and masking of parasitic streaks, elimination of unwanted powder diffraction (e.g., from ice or salt crystals), and pinpointing useful Bragg peaks in each diffraction pattern. The library is equipped with a user-friendly graphical interface for facile parameter adjustment tailored to specific datasets. Compatible with popular SX processing software like OnDA, Cheetah, CrystFEL, and Merge3D, the FDIP library enhances the capabilities of these tools for streamlined and precise serial crystallography analyses.Publicationtype: Journal Article30 100 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: without files , Method and a system for examining sample particles by X-ray radiation(2022-11-09); ; ; ; The invention relates to a method and a system (1) for examining sample particles (2) by X-ray radiation. The method comprises the following steps: a) Generating a flow of a sample fluid (4) along a fluid path (6) in a transport direction Y, wherein sample particles (2) are distributed in the sample fluid (4); b) Exposing the sample fluid (4) flowing through a first application section (8) of the fluid path (6) to a first laser light generated by a first laser light source (38) for a first holding time period such that a first optical light pressure is applied on sample particles (2) entering the first application section (8) resulting in a bunch (12) of sample particles (2), which is released at the end of the first holding time period; c) Repeating step b) such that the fluid flow of the sample fluid (4) leaving the first application section (8) carries a first sequence (14) of spaced bunches (12) of sample particles (2); d) Exposing the sample fluid (4) flowing through a detection section (16) of the fluid path (6) downstream to the first application section (8) to an X-ray set of at least one X-ray pulse generated by an X-ray source (18); e) Repeating step d) to form a second sequence of spaced X-ray sets such that each of the spaced X-ray sets partly or fully intersects a different bunch (12) of the bunches (12) of sample particles (2) carried by the sample fluid (4) flowing through the detection section (16); and f) Detecting by means of an X-ray detector (22) an X-ray radiation formed by the X-ray pulses of the second sequence of spaced X-ray set modified by the bunches (12,32) of sample particles (2) in step e).Publicationtype: Patent29 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: without files , Optical bunching of particles in a liquid flow(2021-10-11); ; ; ; High-speed liquid micro-jets are used to rapidly and repeatedly deliver protein microcrystals to focused and pulsed X-ray beams in the method of serial femtosecond crystallography. However, the current continuous flow of crystals is mismatched to the arrival of X-ray pulses, wasting vast amounts of an often rare and precious sample. Here, we introduce a method to address this problem by periodically trapping and releasing crystals in the liquid flow, creating locally concentrated crystal bunches, using an optical trap integrated in the microfluidic supply line. We experimentally demonstrate a 30-fold increase of particle concentration into 10 Hz bunches of 6.4 µm diameter polystyrene particles. Furthermore, using particle trajectory simulations, a comprehensive description of the optical bunching process and parameter space is presented. Adding this compact optofluidics device to existing injection systems would thereby dramatically reduce sample consumption and extend the application of serial crystallography to a greater range of protein crystal systems that cannot be produced in high abundance. Our approach is suitable for other microfluidic systems that require synchronous measurements of flowing objects.Publicationtype: Journal Article64 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , X-ray screening identifies active site and allosteric inhibitors of SARS-CoV-2 main protease(American Association for the Advancement of Science (Assoc.), 2021-04-02); ; ; ; The coronavirus disease (COVID-19) caused by SARS-CoV-2 is creating tremendous human suffering. To date, no effective drug is available to directly treat the disease. In a search for a drug against COVID-19, we have performed a high-throughput X-ray crystallographic screen of two repurposing drug libraries against the SARS-CoV-2 main protease (Mpro), which is essential for viral replication. In contrast to commonly applied X-ray fragment screening experiments with molecules of low complexity, our screen tested alreadyapproved drugs and drugs in clinical trials. From the three-dimensional protein structures, we identified 37 compounds that bind to Mpro. In subsequent cell-based viral reduction assays, one peptidomimetic and six nonpeptidic compounds showed antiviral activity at nontoxic concentrations. We identified two allosteric binding sites representing attractive targets for drug development against SARS-CoV-2.Publicationtype: Journal ArticleScopus© Citations 214 186 281 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: without files , On the use of multilayer Laue lenses with X-ray free electron lasers(SPIE, 2021); ; We report on the use of multilayer Laue lenses to focus the intense X-ray Free Electron Laser (XFEL) beam at the European XFEL to a spot size of a few tens of nanometers. We present the procedure to align and characterize these lenses and discuss challenges working with the pulse trains from this unique X-ray source.Publicationtype: Conference Paper18 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Segmented flow generator for serial crystallography at the European X-ray free electron laser(2020-09-09); ; ; ; 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.Publicationtype: Journal ArticleScopus© Citations 28 279 184 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , pinkIndexer – a universal indexer for pink-beam X-ray and electron diffraction snapshotsA crystallographic indexing algorithm,pinkIndexer, is presented for the analysisof snapshot diffraction patterns. It can be used in a variety of contexts includingmeasurements made with a monochromatic radiation source, a polychromaticsource or with radiation of very short wavelength. As such, the algorithm isparticularly suited to automated data processing for two emerging measurementtechniques for macromolecular structure determination: serial pink-beam X-raycrystallography and serial electron crystallography, which until now lackedreliable programs for analyzing many individual diffraction patterns fromcrystals of uncorrelated orientation. The algorithm requires approximateknowledge of the unit-cell parameters of the crystal, but not the wavelengthsassociated with each Bragg spot. The use ofpinkIndexeris demonstrated byobtaining 1005 lattices from a published pink-beam serial crystallography dataset that had previously yielded 140 indexed lattices. Additionally, in tests onexperimental serial crystallography diffraction data recorded with quasi-monochromatic X-rays and with electrons the algorithm indexed more patternsthan other programs tested.Publicationtype: Journal ArticleScopus© Citations 24 282 215 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: without files , Time-resolved serial femtosecond crystallography at the European XFEL(2020-01-01); ; ; ; 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.Publicationtype: Journal Article190