搜索结果：crystallography

Customized beam shaping has a wide range of applications from visible light to hard X-rays. While laser beam shaping has matured over recent decades, enabling breakthroughs in optical communication, optical tweezers, and advanced microscopy, extending these techniques to high-brightness X-ray sources could significantly enhance synchrotron applications such as macromolecular crystallography, spectroscopy, and high-resolution imaging. However, X-ray beam shaping remains challenging due to limitations in the available optics and the finite phase-space of synchrotron sources. We introduce a novel method that exploits the monochromatic angular spectrum of undulator radiation combined with the compound refractive lenses (CRLs) to produce a variable circular focal spot with a top-hat intensity profile. By fine-tuning the undulator gap and monochromator settings, this approach enables dynamic control of the spatial beam profile while preserving continuous energy tunability within the limits imposed by the optical configuration and experimental conditions. This technique delivers flexible beam shaping without requiring complex new optical designs, construction, or operational overhead. This method has been successfully demonstrated on a macromolecular crystallography beamline at the Diamond Light Source (DLS), confirming its practicality, adaptability, and potential for widespread adoption in synchrotron-based research.

Aquaporins (AQPs) are membrane channel proteins that facilitate the transport of water and related solutes, playing indispensable roles across a wide range of organisms. Extensive structural studies using X-ray crystallography and electron crystallography have provided fundamental insights into the physiological functions of various AQPs. Recent advances in the structural analysis using single-particle cryo-electron microscopy, which bypasses the requirement for crystallization, have profoundly enhanced our understanding of AQP mechanisms. In this review, we summarize recent progress in AQP biology, providing a comparative analysis of orthodox, glycerol-permeable and unorthodox AQPs.

The inversion of large-scale diffraction datasets from modern synchrotron sources presents a fundamental challenge in computational crystallography. This paper presents a unified algorithmic framework for the analysis of both near-field (morphological) and far-field (orientational and strain) high-energy diffraction microscopy (HEDM) data. We detail the mathematical formalisms and physical models that form the foundation of this methodology. Key aspects include a generalized model for detector distortion correction, robust algorithms for peak identification in noisy and overlapping patterns, a computationally efficient indexing formalism based on Friedel pair symmetry, and a decoupled iterative refinement scheme that exploits the differing sensitivities of position, orientation and lattice parameters to diffraction observables. We also describe the synergistic integration of near-field and far-field data streams, a critical feature of a truly comprehensive approach. The framework is validated in Part II of this series [Sharma et al. (2026). Acta Cryst. A82, https://doi.org/10.1107/S2053273326004018] using both experimental Ti-7 Al datasets and synthetic reconstructions with known ground truth, achieving orientation accuracy of ∼0.05° and position accuracy of ∼10 µm on experimental data, and a 190× improvement in lattice parameter precision over conventional simultaneous parameter refinement on synthetic data. This integrated framework provides a powerful and extensible solution for turning raw diffraction images into actionable microstructural and micromechanical information.

Chemical pollution is a global threat to human health, yet the toxicity mechanisms of most contaminants remains unknown. Here, we applied an ultrahigh-throughput affinity selection-mass spectrometry (AS-MS) platform to systematically identify protein targets of prioritized chemical contaminants. After benchmarking the platform, we screened 50 human proteins against 481 prioritized chemicals, including 446 ToxCast chemicals and 35 per- and polyfluoroalkyl substances (PFAS). Among 24,050 interactions assessed, we discovered 35 interactions involving 13 proteins, with fatty acid-binding proteins (FABPs) emerging as the most ligandable protein family. Given this, we selected FABPs for further validation, which revealed a distinct PFAS binding pattern: legacy PFAS selectively bound to FABP1, whereas replacement compounds, perfluoroether carboxylic acids, unexpectedly interacted with all FABPs. X-ray crystallography further revealed that the ether group enhances the molecular flexibility of alternative PFAS to accommodate the binding pockets of FABPs. Our findings demonstrate that AS-MS is a robust platform for the discovery of protein targets beyond the scope of ToxCast and highlight the broader protein-binding spectrum of alternative PFAS as potential regrettable substitutes.

Respiratory syncytial virus infection (RSV) is a major global health concern, particularly in infants and elderly populations. In this work, we have screened and identified 3 double-stapled peptides derived from a minimal domain of the RSV F heptad repeat, namely 3/4i, 3/4m, and 4/4g, which are potent inhibitors of RSV fusion and remain active against viral escape mutants resistant to small-molecule fusion inhibitors. Our structural activity relationship (SAR) analysis demonstrates that combining a limited set of staples is sufficient to achieve high antiviral potency. X-ray crystallography revealed that the enhanced potency of 3/4i and 3/4m primarily arises from strong hydrophobic interactions between the N-terminal staple and the trimeric HR1 coiled coil of RSV F. In vivo pharmacokinetic, imaging, and feasibility studies in RSV-infected Balb/c mice further support intranasal administration as a promising route for delivering these stapled peptides to the lung, highlighting their potential as therapeutics against RSV.

Advances in structural biology have improved our understanding of the relationship between protein structure and function, while also confirming a widely applicable principle: protein domains with highly conserved three-dimensional folds can perform radically disparate biochemical functions. To gain insight into this structural enigma, we mapped the energetic landscapes of a family of bacterial transcription factors and their anciently diverged structural homologues, the periplasmic binding proteins. Using hydrogen exchange-mass spectrometry, bioinformatics, X-ray crystallography and molecular dynamics, we uncovered an unexpected contrast: despite binding the same sugars, the two families have evolved unique 'energetic blueprints' to support their distinct functional requirements. To test if differences in ensemble energies have functional consequences, we rationally redesigned the protein fold for tunable ligand-driven transcriptional responses. Strikingly, energy-driven protein engineering produced synthetic transcription factors with the theoretically anticipated ligand-induced transcriptional outputs. Thus, decoding energetic blueprints among conserved protein folds provides diverse functional adaptations, paves an alternative roadmap for protein design, and offers a distinct approach for engineering challenging drug targets.

Ferritin-catalysed Fe2+ oxidation by reaction with O2 occurs at an intra-subunit diiron site known as the ferroxidase centre (FoC). Currently, how Fe3+, the key substrate for iron core nucleation/mineralisation, transfers from the FoC to the inner protein surface/central cavity where the mineral is laid down is unknown. Iron-binding sites that become occupied following exposure of anaerobic, Fe2+-bound human cytosolic H-chain ferritin (HuHF) to O2 were identified by time-resolved x-ray crystallography. In addition to the two FoC iron sites, three further sites were identified, each involving Glu61 as a coordinating residue. Substitution by a non-coordinating residue (variant E61A) eliminated binding at these additional iron sites. Solution kinetic studies of Fe2+ oxidation and iron core mineralisation in wild-type HuHF and its E61A variant showed that rapid Fe2+ oxidation was unaffected by loss of Glu61, ruling out an important role for these sites in either guiding Fe2+ to the FoC, or in the mechanism of FoC-catalysed Fe2+ oxidation. Conversely, the transfer of Fe3+ out of the FoC and core mineralisation were both severely affected in the E61A variant. A mechanism for Fe3+ transfer from the FoC to the inner protein surface is proposed.

The European honey bee (Apis mellifera L.) is an essential crop pollinator and is frequently exposed to pesticide residues that may compromise bee health. Mechanisms underlying chemical adaptation and detoxification in honey bees remain incompletely understood, particularly those involving glutathione S-transferases (GSTs). Here, we structurally and functionally characterized omega-class GST AmGSTO1. AmGSTO1 was highly expressed in the fat bodies of nurse and forager bees. X-ray crystallography resolved the glutathione (GSH)-bound AmGSTO1 structure, revealing an active-site cysteine characteristic of omega GSTs. Enzyme assays showed greater catalytic efficiency toward the thioltransferase substrate HED than toward CDNB or PNA. Disc diffusion and bacterial survival assays demonstrated antioxidant activity against cumene hydroperoxide, hydrogen peroxide, and paraquat. Fluorescence binding assays indicated agrochemical binding, while HPLC detected no significant substrate depletion, suggesting a sequestration rather than catalytic role. Overall, AmGSTO1 may contribute to the protection against agrochemical toxicity and oxidative stress in honey bees.

Histone deacetylase 6 (HDAC6) is a cytoplasmic enzyme that deacetylates non-histone substrates such as α-tubulin and cortactin. HDAC6 contains two catalytic domains, each containing a catalytic zinc ion, and a zinc-finger ubiquitin-binding domain. We have discovered BAS-2, a selective HDAC6 inhibitor with an isothiouronium core and no obvious zinc-binding group. To define its mechanism, we combine X-ray crystallography, structure-activity-relationships, molecular modeling and mutagenesis. BAS-2 potently inhibits human HDAC6 but it does not inhibit zebrafish HDAC6. Computational modeling highlighted Asp567 in human HDAC6 as critical for BAS-2 recognition and mutational analyses confirmed this. The corresponding zebrafish residue is Asn530 and the crystal structure of the N530D variant zHDAC6 revealed binding of a BAS-2-derived mercaptoacetamide that engages the catalytic zinc via strong thiolate-zinc coordination. Leveraging the orientation of BAS-2 binding, we designed a BAS-2-based proteolysis targeting chimera that induced proteasome-dependent HDAC6 degradation in cells, verified by global proteomics. Collectively, these insights clarify species selectivity and demonstrate that BAS-2 acts as a selective, mechanism-based inhibitor of human HDAC6. These discoveries will aid the development of the next generation of selective HDAC6 inhibitors and degraders.

π-Conjugated compounds containing tricoordinate boron atoms have attracted significant attention because of their unique properties, including low-lying LUMO energy levels, extended π-conjugation, and high Lewis acidity. In particular, planarized triarylboranes have been extensively studied in recent years owing to their high chemical stability and excellent photoluminescence properties. Despite exhibiting potential for strong intermolecular interactions and superior semiconducting performance arising from the absence of bulky substituents, the synthesis of p-π* conjugated polymers using such planarized units as building blocks, along with the evaluation of their semiconducting properties, remains a significant challenge that has not yet been addressed. In this study, we report the synthesis of DTTB, a new planarized triarylborane featuring benzene and thiophene rings fused via boron and sulfur atoms. DTTB represents a versatile building block that combines good chemical stability with a perfectly planar geometry, facilitating intermolecular π-π stacking as confirmed by X-ray crystallography. Furthermore, the fused thiophene rings allowed for facile deprotonation at the α-positions, enabling the first integration of planarized triarylborane units into p-π* conjugated polymers. The synthesized p-π* conjugated polymers exhibited ambipolar transport behavior with enhanced carrier mobilities compared to non-planarized analogs. This work provides a conceptual advance by demonstrating that the rigid planarization of triarylborane units is a superior strategy for enhancing the semiconducting performance and thermal stability of p-π* conjugated materials, offering a design strategy for polymer semiconductors.

Molecular catalysts built from earth-abundant metals for hydrogen evolution reaction (HER) often struggle to combine high activity, extended lifetimes, and well-defined mechanisms. We reported three nickel(II) complexes, [{dmit(Py')2}NiX2] (where X is Cl (1), Br (2), and NCS (3), while dmit(Py')2 is 4,5-bis((3,4-dimethoxypyridin-2-yl)methylthio)-1,3-dithiole-2-thione), that share an S2N2 ligand framework integrating proton-relaying pyridyl groups into the redox-active dmit backbone. X-ray crystallography confirmed a distorted octahedral geometry at nickel. These complexes catalyzed proton reduction with overpotentials (η) of 0.67-0.79 V in 53.80 mM trifluoroacetic acid (TFA), as determined by cyclic voltammetry. Controlled potential electrolysis (CPE) of complex 2 for 8 h achieved a turnover number (TON) of 16.8 with 85.61% Faradaic efficiency. An inverse kinetic isotope effect (KIE = 0.70-0.74) suggested the formation of Ni-H intermediates, while DFT calculations supported a ligand-assisted, metal-centered ECEC mechanism. UV-vis monitoring, postelectrolysis voltammetry, and SEM imaging all pointed to homogeneous catalysis and robust complex stability. These results demonstrate that embedding both proton-shuttling sites and redox noninnocence within the ligand structure enables the development of high-performance HER catalysts, providing a rational design framework for bioinspired catalysts.

A pair of isomeric sesquiterpenes, alantolactone and isoalantolactone, were isolated from Inula racemosa Hook. f. and utilised for the synthesis of diverse natural product hybrids using azomethine ylide cycloaddition. In this particular study, we employed benzylamine and diketones (isatin, ninhydrin, and acenaphthoquinone) to generate the azomethine ylide in situ. The azomethine ylide reacts in a chemo- and regioselective manner with the active exocyclic double bond of alantolactone and isoalantolactone to produce a library of Twelve (12) dispiro hybrids. All synthesised compounds were extensively characterised using 1D and 2D NMR spectroscopy and X-ray crystallography. This is the first report on azomethine ylide cycloaddition using benzylamines and 4-picolylamine for the synthesis of dispiro hybrids of sesquiterpene lactones. The unique structural scaffolds of these compounds may be utilised in drug discovery.

Structure-based drug discovery (SBDD) aims to identify novel molecules that bind to therapeutic protein targets. The vast chemical space and limitations of traditional approaches make this task challenging. Recent advances in AI-generative models, such as flow matching, can produce novel, pocket-conditioned molecular structures directly in three-dimensional space. However, most pocket conditioned models in the literature are trained on structures derived from the Protein Data Bank (PDB), which contains structures with varying quality and inconsistent annotation. Moreover, the PDB is enriched with cofactors and natural products, thereby poorly representing real world SBDD scenarios. The relatively limited number of ligand series within the same pockets also hinder the model's ability to learn protein-ligand interactions effectively. Here for the first time we report the results of training pocket-conditioned generative models on internal crystallography data from a large pharmaceutical company. We also investigate other key determinants of model performance, such as inclusion of hydrogens and pretraining on unconditional data. We evaluate how each factor affects the generative quality of the ligands across the diverse training settings. Our results provide practical guidelines for the development of more effective 3D generative models for SBDD and highlight key directions for future research toward reliable, pocket-aware molecular design.

SixA is a phosphatase that dephosphorylates phosphohistidine in proteins involved in signaling in bacteria. Using NMR spectroscopy, the interaction of SixA was structurally characterized with one of its functional substrates, the phosphocarrier protein NPr, which shuttles a phosphate group between components of the nitrogen-related phosphotransferase system using a phosphorylated histidine residue. An ensemble structure of the complex was generated combining chemical shift perturbation data and intermolecular paramagnetic relaxation enhancement (PRE) via Gd-DOTA spin-labels attached to NPr, which shows two regions of interaction outside the SixA active site. Simulated molecular crowding did not enhance interaction with these regions. Phosphate, one of the products of the enzyme catalysis, was determined to bind at a site consistent with that seen for a tungstate ion in the SixA X-ray crystallography structure. SixA dephosphorylates NPr too rapidly for NMR characterization of the phosphorylated form; consequently, interaction with substrate analogs was studied, including a mutant of NPr, H16D, to mimic the negative charge of phosphate at the histidine, and non-cleavable analogs of phosphohistidine. The non-cleavable analogs bound in the active site, but at a location distinct from bound phosphate. A phosphohistidine-containing tripeptide was synthesized, and the activity of SixA was measured using liquid chromatography/mass spectrometry. In the presence of phosphate, the activity of SixA was enhanced.

Zn-based aqueous batteries have attracted widespread research attention, while the lack of nucleation theory for electrochemical interactions at the Zn-water interface constrains efforts to suppress the thermodynamically spontaneous hydrogen evolution reaction and dendrite formation, thereby stalling practical development. Elucidating Zn electrodeposition in aqueous media requires Zn-specific nucleation theory and a descriptor to regulate interfacial electrochemistry. Conventional Li-based spherical nucleation models disregard Zn's crystallography and the interfacial resistance that governs nucleation, thereby focusing on polarization variations. In this work, we reformulate the classical spherical nucleation theory derived by Li for the hexagonal close-packed structure of Zn and establish a dimensionless descriptor (Wf) to quantitatively rationalize interfacial electrochemistry. Wf synthesizes the polarization driving force and interfacial resistance into a stability metric. Higher Wf values facilitate uniform Zn deposition, as evidenced by the literature. Accordingly, we develop a high-Wf electrolyte to inhibit dendrites and side reactions, achieving over 700 h at 100% depth of discharge and 7660 cycles at 10 A g-1 in a Zn||NaV3O8 cell. This work provides a fundamental nucleation theory and a generally applicable quantitative metric for the rational design of Zn-based aqueous batteries.