Medical contrast agents as promising tools for biomacromolecular SAXS experiments
Small-angle X-ray scattering (SAXS) has become an indispensable tool in structural biology, complementing atomic-resolution techniques. It is sensitive to the electron-density difference between solubilized biomacromolecules and the buffer, and provides information on molecular masses, particle dimensions and interactions, low-resolution conformations and pair distance-distribution functions. When SAXS data are recorded at multiple contrasts, i.e. at different solvent electron densities, it is possible to probe, in addition to their overall shape, the internal electron-density profile of biomacromolecular assemblies. Unfor...
Source: Acta Crystallographica Section D - August 9, 2022 Category: Biochemistry Authors: Gabel, F. Engilberge, S. Schmitt, E. Thureau, A. Mechulam, Y. P é rez, J. Girard, E. Tags: small-angle scattering SAXS contrast variation macromolecular complexes electron density medical contrast agents iohexol Gd-HPDO3A tools for SAXS research papers Source Type: research
Structural basis for the acetylation mechanism of the Legionella effector VipF
The pathogen Legionella pneumophila, which is the causative agent of Legionnaires' disease, secrets hundreds of effectors into host cells via its Dot/Icm secretion system to subvert host-cell pathways during pathogenesis. VipF, a conserved core effector among Legionella species, is a putative acetyltransferase, but its structure and catalytic mechanism remain unknown. Here, three crystal structures of VipF in complex with its cofactor acetyl-CoA and/or a substrate are reported. The two GNAT-like domains of VipF are connected as two wings by two β -strands to form a U-shape. Both domains bind acetyl-CoA or CoA, but only in...
Source: Acta Crystallographica Section D - August 9, 2022 Category: Biochemistry Authors: Chen, T.-T. Lin, Y. Zhang, S. Han, A. Tags: Legionella pneumophila Legionnaires' disease core effectors acetyltransferases VipF research papers Source Type: research
Native glycosylation and binding of the antidepressant paroxetine in a low-resolution crystal structure of human myeloperoxidase
Human myeloperoxidase (MPO) utilizes hydrogen peroxide to oxidize organic compounds and as such plays an essential role in cell-component synthesis, in metabolic and elimination pathways, and in the front-line defence against pathogens. Moreover, MPO is increasingly being reported to play a role in inflammation. The enzymatic activity of MPO has also been shown to depend on its glycosylation. Mammalian MPO crystal structures deposited in the Protein Data Bank (PDB) present only a partial identification of their glycosylation. Here, a newly obtained crystal structure of MPO containing four disulfide-linked dimers and showin...
Source: Acta Crystallographica Section D - August 9, 2022 Category: Biochemistry Authors: Krawczyk, L. Semwal, S. Soubhye, J. Lemri Ouadriri, S. Pr é vost, M. Van Antwerpen, P. Roos, G. Bouckaert, J. Tags: human myeloperoxidase glycosylation N-glycan refinement thiocyanate paroxetine feature articles Source Type: research
The crystal structure of CbpD clarifies substrate-specificity motifs in chitin-active lytic polysaccharide monooxygenases
Pseudomonas aeruginosa secretes diverse proteins via its type 2 secretion system, including a 39 kDa chitin-binding protein, CbpD. CbpD has recently been shown to be a lytic polysaccharide monooxygenase active on chitin and to contribute substantially to virulence. To date, no structure of this virulence factor has been reported. Its first two domains are homologous to those found in the crystal structure of Vibrio cholerae GbpA, while the third domain is homologous to the NMR structure of the CBM73 domain of Cellvibrio japonicus CjLPMO10A. Here, the 3.0 Å resolution crystal structure of CbpD solved by molecular r...
Source: Acta Crystallographica Section D - July 27, 2022 Category: Biochemistry Authors: Dade, C.M. Douzi, B. Cambillau, C. Ball, G. Voulhoux, R. Forest, K.T. Tags: LPMOs lytic polysaccharide monooxygenases AA10 Pseudomonas aeruginosa chitin binding protein CbpD type 2 secretion systems RoseTTAFold biofuels bacterial pathogenesis research papers Source Type: research
α -SAS: an integrative approach for structural modeling of biological macromolecules in solution
Modern small-angle scattering (SAS) experiments with neutrons (SANS) or X-rays (SAXS) combined with contrast variation provide comprehensive information about the structure of large multicomponent macromolecules in solution and allow the size, shape and relative arrangement of each component to be mapped out. To obtain such information, it is essential to perform well designed experiments, in which all necessary steps, from assessing sample suitability to structure modeling, are properly executed. This paper describes α -SAS, an integrative approach that is useful for effectively planning a biological contrast-variation S...
Source: Acta Crystallographica Section D - July 27, 2022 Category: Biochemistry Authors: Anitas, E.M. Tags: small-angle neutron scattering SAS contrast variation Monte Carlo simulations biological macromolecules α -shapes research papers Source Type: research
Native SAD phasing at room temperature
Single-wavelength anomalous diffraction (SAD) is a routine method for overcoming the phase problem when solving macromolecular structures. This technique requires the accurate measurement of intensities to determine differences between Bijvoet pairs. Although SAD experiments are commonly conducted at cryogenic temperatures to mitigate the effects of radiation damage, such temperatures can alter the conformational ensemble of the protein and may impede the merging of data from multiple crystals due to non-uniform freezing. Here, a strategy is presented to obtain high-quality data from room-temperature, single-crystal experi...
Source: Acta Crystallographica Section D - July 27, 2022 Category: Biochemistry Authors: Greisman, J.B. Dalton, K.M. Sheehan, C.J. Klureza, M.A. Kurinov, I. Hekstra, D.R. Tags: X-ray crystallography native SAD room temperature phasing model building research papers Source Type: research
Room-temperature serial synchrotron crystallography of Drosophila cryptochrome
Fixed-target serial crystallography allows the high-throughput collection of diffraction data from small crystals at room temperature. This methodology is particularly useful for difficult samples that have sensitivity to radiation damage or intolerance to cryoprotection measures; fixed-target methods also have the added benefit of low sample consumption. Here, this method is applied to the structure determination of the circadian photoreceptor cryptochrome (CRY), previous structures of which have been determined at cryogenic temperature. In determining the structure, several data-filtering strategies were tested for combi...
Source: Acta Crystallographica Section D - July 27, 2022 Category: Biochemistry Authors: Schneps, C.M. Ganguly, A. Crane, B.R. Tags: circadian clock cryptochromes circadian photoreceptor cryptochrome flavin adenine dinucleotide fixed-target serial synchrotron crystallography protein crystallography correlation filtering B factors molecular dynamics room temperature Source Type: research
Probing ligand binding of endothiapepsin by `temperature-resolved' macromolecular crystallography
Continuous developments in cryogenic X-ray crystallography have provided most of our knowledge of 3D protein structures, which has recently been further augmented by revolutionary advances in cryoEM. However, a single structural conformation identified at cryogenic temperatures may introduce a fictitious structure as a result of cryogenic cooling artefacts, limiting the overview of inherent protein physiological dynamics, which play a critical role in the biological functions of proteins. Here, a room-temperature X-ray crystallographic method using temperature as a trigger to record movie-like structural snapshots has been...
Source: Acta Crystallographica Section D - July 27, 2022 Category: Biochemistry Authors: Huang, C.-Y. Aumonier, S. Engilberge, S. Eris, D. Smith, K.M.L. Leonarski, F. Wojdyla, J.A. Beale, J.H. Buntschu, D. Pauluhn, A. Sharpe, M.E. Metz, A. Olieric, V. Wang, M. Tags: conformational heterogeneity protein plasticity fragment binding endothiapepsin room temperature macromolecular crystallography research papers Source Type: research
Knowledge-based prediction of DNA hydration using hydrated dinucleotides as building blocks
Water plays an important role in stabilizing the structure of DNA and mediating its interactions. Here, the hydration of DNA was analyzed in terms of dinucleotide fragments from an ensemble of 2727 nonredundant DNA chains containing 41 853 dinucleotides and 316 265 associated first-shell water molecules. The dinucleotides were classified into categories based on their 16 sequences and the previously determined structural classes known as nucleotide conformers (NtCs). The construction of hydrated dinucleotide building blocks allowed dinucleotide hydration to be calculated as the probability of water density distribu...
Source: Acta Crystallographica Section D - July 21, 2022 Category: Biochemistry Authors: Biedermannov á , L. Č ern ý , J. Mal ý , M. Nekardov á , M. Schneider, B. Tags: DNA hydration water dinucleotide fragments knowledge-based prediction WatNA research papers Source Type: research
Data collection from crystals grown in microfluidic droplets
Protein crystals grown in microfluidic droplets have been shown to be an effective and robust platform for storage, transport and serial crystallography data collection with a minimal impact on diffraction quality. Single macromolecular microcrystals grown in nanolitre-sized droplets allow the very efficient use of protein samples and can produce large quantities of high-quality samples for data collection. However, there are challenges not only in growing crystals in microfluidic droplets, but also in delivering the droplets into X-ray beams, including the physical arrangement, beamline and timing constraints and ease of ...
Source: Acta Crystallographica Section D - July 21, 2022 Category: Biochemistry Authors: Babnigg, G. Sherrell, D. Kim, Y. Johnson, J.L. Nocek, B. Tan, K. Axford, D. Li, H. Bigelow, L. Welk, L. Endres, M. Owen, R.L. Joachimiak, A. Tags: X-ray data collection fixed targets serial crystallography sialate O-acetylesterase fucosidases α – β fold hydrolases microfluidic droplets low dose emulsions research papers Source Type: research
Structural dynamics: review of time-resolved cryo-EM
The structural determination of biological macromolecules has been transformative for understanding biochemical mechanisms and developing therapeutics. However, the ultimate goal of characterizing how structural dynamics underpin biochemical processes has been difficult. This is largely due to significant technical challenges that hinder data collection and analysis on the native timescales of macromolecular dynamics. Single-particle cryo-EM provides a powerful platform to approach this challenge, since samples can be frozen faster than the single-turnover timescales of most biochemical reactions. In order to enable time-r...
Source: Acta Crystallographica Section D - July 21, 2022 Category: Biochemistry Authors: M ä eots, M.-E. Enchev, R.I. Tags: cryo-EM time-resolved cryo-EM sample preparation structural biology structural dynamics topical reviews Source Type: research
Application of sulfur SAD to small crystals with a large asymmetric unit and anomalous substructure
The application of sulfur single-wavelength anomalous dispersion (S-SAD) to determine the crystal structures of macromolecules can be challenging if the asymmetric unit is large, the crystals are small, the size of the anomalously scattering sulfur structure is large and the resolution at which the anomalous signals can be accurately measured is modest. Here, as a study of such a case, approaches to the SAD phasing of orthorhombic Ric-8A crystals are described. The structure of Ric-8A was published with only a brief description of the phasing process [Zeng et al. (2019), Structure, 27, 1137 – 1141]. Here, alternative app...
Source: Acta Crystallographica Section D - July 14, 2022 Category: Biochemistry Authors: Mou, T.-C. Zeng, B. Doukov, T.I. Sprang, S.R. Tags: sulfur SAD phasing data scaling single-wavelength anomalous dispersion sulfur substructure determination large asymmetric unit Ric-8A research papers Source Type: research
Theoretical 3D electron diffraction electrostatic potential maps of proteins modeled with a multipolar pseudoatom data bank
The availability of atomic resolution experimental maps of electrostatic potential from 3D electron diffraction (3D ED) extends the possibility of investigating the electrostatic potential beyond the determination of non-H-atom positions. However, accurate tools to calculate this potential for macromolecules, without the use of expensive quantum calculations, are lacking. The University at Buffalo Data Bank (UBDB) gathers atom types that can be used to calculate accurate electrostatic potential maps via structure-factor calculations. Here, the transferable aspherical atom model (TAAM) is applied with UBDB to investigate th...
Source: Acta Crystallographica Section D - July 14, 2022 Category: Biochemistry Authors: Kulik, M. Chodkiewicz, M.L. Dominiak, P.M. Tags: electron diffraction transferable aspherical atom model multipolar scattering factors cryo-electron microscopy quantum crystallography electrostatic potential maps UBDB macromolecular structure proteins research papers Source Type: research
Evaluating the impact of X-ray damage on conformational heterogeneity in room-temperature (277 K) and cryo-cooled protein crystals
Cryo-cooling has been nearly universally adopted to mitigate X-ray damage and facilitate crystal handling in protein X-ray crystallography. However, cryo X-ray crystallographic data provide an incomplete window into the ensemble of conformations that is at the heart of protein function and energetics. Room-temperature (RT) X-ray crystallography provides accurate ensemble information, and recent developments allow conformational heterogeneity (the experimental manifestation of ensembles) to be extracted from single-crystal data. Nevertheless, high sensitivity to X-ray damage at RT raises concerns about data reliability. To ...
Source: Acta Crystallographica Section D - July 14, 2022 Category: Biochemistry Authors: Yabukarski, F. Doukov, T. Mokhtari, D.A. Du, S. Herschlag, D. Tags: protein X-ray crystallography X-ray damage room temperature conformational ensembles hydrogen bonds research papers Source Type: research
Training undergraduate research assistants with an outcome-oriented and skill-based mentoring strategy
This report especially shows how the Critical Assessment of Protein Structure Prediction (CASP14) challenge was used to amplify student research efforts. As a result of this challenge, multiple publications, presentations and scholarships were awarded to the participating students. The mentoring framework continues to see much success in allowing undergraduate students, including students from underrepresented groups, to foster scientific talent and make meaningful contributions to the scientific community. (Source: Acta Crystallographica Section D)
Source: Acta Crystallographica Section D - July 14, 2022 Category: Biochemistry Authors: Della Corte, D. Morris, C.J. Billings, W.M. Stern, J. Jarrett, A.J. Hedelius, B. Bennion, A. Tags: mentoring undergraduates CASP protein structure prediction deep learning research papers Source Type: research
