STOPGAP: an open-source package for template matching, subtomogram alignment and classification
Cryo-electron tomography (cryo-ET) enables molecular-resolution 3D imaging of complex biological specimens such as viral particles, cellular sections and, in some cases, whole cells. This enables the structural characterization of molecules in their near-native environments, without the need for purification or separation, thereby preserving biological information such as conformational states and spatial relationships between different molecular species. Subtomogram averaging is an image-processing workflow that allows users to leverage cryo-ET data to identify and localize target molecules, determine high-resolution stru...
Source: Acta Crystallographica Section D - April 12, 2024 Category: Biochemistry Authors: Wan, W. Khavnekar, S. Wagner, J. Tags: STOPGAP cryo-ET open source software subtomogram alignment research papers Source Type: research
HEIDI: an experiment-management platform enabling high-throughput fragment and compound screening
The Swiss Light Source facilitates fragment-based drug-discovery campaigns for academic and industrial users through the Fast Fragment and Compound Screening (FFCS) software suite. This framework is further enriched by the option to utilize the Smart Digital User (SDU) software for automated data collection across the PXI, PXII and PXIII beamlines. In this work, the newly developed HEIDI webpage (https://heidi.psi.ch) is introduced: a platform crafted using state-of-the-art software architecture and web technologies for sample management of rotational data experiments. The HEIDI webpage features a data-review tab for en...
Source: Acta Crystallographica Section D - April 12, 2024 Category: Biochemistry Authors: Metz, A. Stegmann, D.P. Panepucci, E.H. Buehlmann, S. Huang, C.-Y. McAuley, K.E. Wang, M. Wojdyla, J.A. Sharpe, M.E. Smith, K.M.L. Tags: HEIDI FFCS software suite data management fragment screening high-throughput crystallography research papers Source Type: research
Scaling and merging macromolecular diffuse scattering with mdx2
Diffuse scattering is a promising method to gain additional insight into protein dynamics from macromolecular crystallography experiments. Bragg intensities yield the average electron density, while the diffuse scattering can be processed to obtain a three-dimensional reciprocal-space map that is further analyzed to determine correlated motion. To make diffuse scattering techniques more accessible, software for data processing called mdx2 has been created that is both convenient to use and simple to extend and modify. mdx2 is written in Python, and it interfaces with DIALS to implement self-contained data-reduction work...
Source: Acta Crystallographica Section D - April 12, 2024 Category: Biochemistry Authors: Meisburger, S.P. Ando, N. Tags: mdx2 diffuse scattering macromolecular crystallography data processing software reciprocal-space mapping research papers Source Type: research
EMinsight: a tool to capture cryoEM microscope configuration and experimental outcomes for analysis and deposition
The widespread adoption of cryoEM technologies for structural biology has pushed the discipline to new frontiers. A significant worldwide effort has refined the single-particle analysis (SPA) workflow into a reasonably standardized procedure. Significant investments of development time have been made, particularly in sample preparation, microscope data-collection efficiency, pipeline analyses and data archiving. The widespread adoption of specific commercial microscopes, software for controlling them and best practices developed at facilities worldwide has also begun to establish a degree of standardization to data structu...
Source: Acta Crystallographica Section D - March 26, 2024 Category: Biochemistry Authors: Hatton, D. Cha, J. Riggs, S. Harrison, P.J. Thiyagalingam, J. Clare, D.K. Morris, K.L. Tags: EMinsight cryoEM data mining deposition machine learning research papers Source Type: research
Mononuclear binding and catalytic activity of europium(III) and gadolinium(III) at the active site of the model metalloenzyme phosphotriesterase
Lanthanide ions have ideal chemical properties for catalysis, such as hard Lewis acidity, fast ligand-exchange kinetics, high coordination-number preferences and low geometric requirements for coordination. As a result, many small-molecule lanthanide catalysts have been described in the literature. Yet, despite the ability of enzymes to catalyse highly stereoselective reactions under gentle conditions, very few lanthanoenzymes have been investigated. In this work, the mononuclear binding of europium(III) and gadolinium(III) to the active site of a mutant of the model enzyme phosphotriesterase are described using X-ray crys...
Source: Acta Crystallographica Section D - March 21, 2024 Category: Biochemistry Authors: Breeze, C.W. Nakano, Y. Campbell, E.C. Frkic, R.L. Lupton, D.W. Jackson, C.J. Tags: lanthanides phosphotriesterase anomalous scattering research papers Source Type: research
Tomo Live: an on-the-fly reconstruction pipeline to judge data quality for cryo-electron tomography workflows
Data acquisition and processing for cryo-electron tomography can be a significant bottleneck for users. To simplify and streamline the cryo-ET workflow, Tomo Live, an on-the-fly solution that automates the alignment and reconstruction of tilt-series data, enabling real-time data-quality assessment, has been developed. Through the integration of Tomo Live into the data-acquisition workflow for cryo-ET, motion correction is performed directly after each of the acquired tilt angles. Immediately after the tilt-series acquisition has completed, an unattended tilt-series alignment and reconstruction into a 3D volume is performed...
Source: Acta Crystallographica Section D - March 21, 2024 Category: Biochemistry Authors: Comet, M. Dijkman, P.M. Boer Iwema, R. Franke, T. Masiulis, S. Schampers, R. Raschdorf, O. Grollios, F. Pryor, E.E. Drulyte, I. Tags: electron tomography real-time processing cryo-ET Tomo Live research papers Source Type: research
Efficient in situ screening of and data collection from microcrystals in crystallization plates
A considerable bottleneck in serial crystallography at XFEL and synchrotron sources is the efficient production of large quantities of homogenous, well diffracting microcrystals. Efficient high-throughput screening of batch-grown microcrystals and the determination of ground-state structures from different conditions is thus of considerable value in the early stages of a project. Here, a highly sample-efficient methodology to measure serial crystallography data from microcrystals by raster scanning within standard in situ 96-well crystallization plates is described. Structures were determined from very small quantities of ...
Source: Acta Crystallographica Section D - March 15, 2024 Category: Biochemistry Authors: Thompson, A.J. Sanchez-Weatherby, J. Williams, L.J. Mikolajek, H. Sandy, J. Worrall, J.A.R. Hough, M.A. Tags: serial crystallography in situ data collection peroxidase radiation damage microcrystals research papers Source Type: research
VitroJet: new features and case studies
Single-particle cryo-electron microscopy has become a widely adopted method in structural biology due to many recent technological advances in microscopes, detectors and image processing. Before being able to inspect a biological sample in an electron microscope, it needs to be deposited in a thin layer on a grid and rapidly frozen. The VitroJet was designed with this aim, as well as avoiding the delicate manual handling and transfer steps that occur during the conventional grid-preparation process. Since its creation, numerous technical developments have resulted in a device that is now widely utilized in multiple laborat...
Source: Acta Crystallographica Section D - March 15, 2024 Category: Biochemistry Authors: Henderikx, R.J.M. Mann, D. Domanska, A. Dong, J. Shahzad, S. Lak, B. Filopoulou, A. Ludig, D. Grininger, M. Momoh, J. Laanto, E. Oksanen, H.M. Bisikalo, K. Williams, P.A. Butcher, S.J. Peters, P.J. Beulen, B.W.A.M.M. Tags: cryo-EM VitroJet ice thickness pin printing jet vitrification research papers Source Type: research
AlphaFold-assisted structure determination of a bacterial protein of unknown function using X-ray and electron crystallography
Macromolecular crystallography generally requires the recovery of missing phase information from diffraction data to reconstruct an electron-density map of the crystallized molecule. Most recent structures have been solved using molecular replacement as a phasing method, requiring an a priori structure that is closely related to the target protein to serve as a search model; when no such search model exists, molecular replacement is not possible. New advances in computational machine-learning methods, however, have resulted in major advances in protein structure predictions from sequence information. Methods that generate ...
Source: Acta Crystallographica Section D - March 7, 2024 Category: Biochemistry Authors: Miller, J.E. Agdanowski, M.P. Dolinsky, J.L. Sawaya, M.R. Cascio, D. Rodriguez, J.A. Yeates, T.O, Tags: electron diffraction protein structure prediction AlphaFold bacterial proteins molecular replacement research papers Source Type: research
Structural determination and modeling of ciliary microtubules
The axoneme, a microtubule-based array at the center of every cilium, has been the subject of structural investigations for decades, but only recent advances in cryo-EM and cryo-ET have allowed a molecular-level interpretation of the entire complex to be achieved. The unique properties of the nine doublet microtubules and central pair of singlet microtubules that form the axoneme, including the highly decorated tubulin lattice and the docking of massive axonemal complexes, provide opportunities and challenges for sample preparation, 3D reconstruction and atomic modeling. Here, the approaches used for cryo-EM and cryo-ET...
Source: Acta Crystallographica Section D - March 7, 2024 Category: Biochemistry Authors: Walton, T. Doran, M.H. Brown, A. Tags: cryo-EM cryo-ET cilia axonemes microtubules research papers Source Type: research
Welcoming five new Co-editors
(Source: Acta Crystallographica Section D)
Source: Acta Crystallographica Section D - March 1, 2024 Category: Biochemistry Authors: Bond, C. Garman, E.F. Read, R.J. Tags: Editors structural biology cryoEM electron diffraction editorial Source Type: research
Characterization of novel mevalonate kinases from the tardigrade Ramazzottius varieornatus and the psychrophilic archaeon Methanococcoides burtonii
Mevalonate kinase is central to the isoprenoid biosynthesis pathway. Here, high-resolution X-ray crystal structures of two mevalonate kinases are presented: a eukaryotic protein from Ramazzottius varieornatus and an archaeal protein from Methanococcoides burtonii. Both enzymes possess the highly conserved motifs of the GHMP enzyme superfamily, with notable differences between the two enzymes in the N-terminal part of the structures. Biochemical characterization of the two enzymes revealed major differences in their sensitivity to geranyl pyrophosphate and farnesyl pyrophosphate, and in their thermal stabilities. This work ...
Source: Acta Crystallographica Section D - February 27, 2024 Category: Biochemistry Authors: Esquirol, L. Newman, J. Nebl, T. Scott, C. Vickers, C. Sainsbury, F. Peat, T.S. Tags: mevalonate kinases GHMP enzyme superfamily feedback inhibition Ramazzottius varieornatus Methanococcoides burtonii research papers Source Type: research
XFEL structure of carbonic anhydrase II: a comparative study of XFEL, NMR, X-ray and neutron structures
The combination of X-ray free-electron lasers (XFELs) with serial femtosecond crystallography represents cutting-edge technology in structural biology, allowing the study of enzyme reactions and dynamics in real time through the generation of `molecular movies'. This technology combines short and precise high-energy X-ray exposure to a stream of protein microcrystals. Here, the XFEL structure of carbonic anhydrase II, a ubiquitous enzyme responsible for the interconversion of CO2 and bicarbonate, is reported, and is compared with previously reported NMR and synchrotron X-ray and neutron single-crystal structures. (Source: ...
Source: Acta Crystallographica Section D - February 27, 2024 Category: Biochemistry Authors: Hull, J.A. Lee, C. Kim, J.K. Lim, S.W. Park, J. Park, S. Lee, S.J. Park, G. Eom, I. Kim, M. Hyun, H. Combs, J.E. Andring, J.T. Lomelino, C. Kim, C.U. McKenna, R. Tags: carbonic anhydrase II XFELs X-ray crystallography NMR neutron diffraction crystallography radiation damage research papers Source Type: research
Advanced exploitation of unmerged reflection data during processing and refinement with autoPROC and BUSTER
The validation of structural models obtained by macromolecular X-ray crystallography against experimental diffraction data, whether before deposition into the PDB or after, is typically carried out exclusively against the merged data that are eventually archived along with the atomic coordinates. It is shown here that the availability of unmerged reflection data enables valuable additional analyses to be performed that yield improvements in the final models, and tools are presented to implement them, together with examples of the results to which they give access. The first example is the automatic identification and remov...
Source: Acta Crystallographica Section D - February 27, 2024 Category: Biochemistry Authors: Vonrhein, C. Flensburg, C. Keller, P. Fogh, R. Sharff, A. Tickle, I.J. Bricogne, G. Tags: unmerged reflection data fitness analysis reflection auditing validation against diffraction data radiation damage research papers Source Type: research
A service-based approach to cryoEM facility processing pipelines at eBIC
Electron cryo-microscopy image-processing workflows are typically composed of elements that may, broadly speaking, be categorized as high-throughput workloads which transition to high-performance workloads as preprocessed data are aggregated. The high-throughput elements are of particular importance in the context of live processing, where an optimal response is highly coupled to the temporal profile of the data collection. In other words, each movie should be processed as quickly as possible at the earliest opportunity. The high level of disconnected parallelization in the high-throughput problem directly allows a complet...
Source: Acta Crystallographica Section D - February 20, 2024 Category: Biochemistry Authors: Horstmann, A. Riggs, S. Chaban, Y. Clare, D.K. de Freitas, G. Farmer, D. Howe, A. Morris, K.L. Hatton, D. Tags: cryoEM facilities eBIC processing pipelines cloud computing research papers Source Type: research
