The inherent ambiguity in reconstructed images from coherent diffraction imaging (CDI) poses an intrinsic challenge, as images derived from the same dataset under varying initial conditions often display inconsistencies. This study introduces a method that employs the Noise2Noise approach combined with neural networks to effectively mitigate these ambiguities. We applied this methodology to hundreds of ambiguous reconstructed images retrieved from a single diffraction pattern using a conventional retrieval algorithm. Our results demonstrate that ambiguous features in these reconstructions are effectively treated as inter-reconstruction noise and are significantly reduced. The post-Noise2Noise treated images closely approximate the average and singular value decomposition analysis of various reconstructions, providing consistent and reliable reconstructions.

The crystal structure of a glycosyl­ated porphyrin (P_Gal2) system, C70H70N4O12, where two iso­propyl­idene protected galactose moieties are attached to the meso position of a substituted tetra­aryl porphyrin is reported. This structure reveals that the parent porphyrin is planar, with the galactose moieties positioned above and below the porphyrin macrocycle. This orientation likely prevents porphyrin–porphyrin H-type aggregation, potentially enhancing its efficiency as a photosensitizer in photodynamic therapy. Notable non-bonding C—H⋯O and C—H⋯π inter­actions among adjacent P_Gal2 systems are observed in this crystal network. Additionally, the tolyl groups of each porphyrin can engage in π–π inter­actions with the delocalized π-systems of neighboring porphyrins.

The crystal structure of a new 1:1 cocrystal of carbamazepine and S-naproxen (C15H12N2O·C14H14O3) was solved from powder X-ray diffraction (PXRD). The PXRD pattern was measured at the high-resolution beamline CRISTAL at synchrotron SOLEIL (France). The structure was solved using Monte Carlo simulated annealing, then refined with Rietveld refinement. The positions of the H atoms were obtained from density functional theory (DFT) ground-state calculations. The symmetry is ortho­rhom­bic with the space group P212121 (No. 19) and the following lattice parameters: a = 33.5486 (9), b = 26.4223 (6), c = 5.3651 (10) Å and V = 4755.83 (19) Å3.

Beauveriolides, including the main beauveriolide I {systematic name: (3R,6S,9S,13S)-9-benzyl-13-[(2S)-hexan-2-yl]-6-methyl-3-(2-methyl­prop­yl)-1-oxa-4,7,10-tri­aza­cyclo­tridecane-2,5,8,11-tetrone, C27H41N3O5}, are a series of cyclo­depsipeptides that have shown promising results in the treatment of Alzheimer's disease and in the prevention of foam cell formation in atherosclerosis. Their crystal structure studies have been difficult due to their tiny crystal size and fibre-like morphology, until now. Recent developments in 3D electron diffraction methodology have made it possible to accurately study the crystal structures of submicron crystals by overcoming the problems of beam sensitivity and dynamical scattering. In this study, the absolute structure of beauveriolide I was determined by 3D electron diffraction. The cyclo­dep­si­peptide crystallizes in the space group I2 with lattice parameters a = 40.2744 (4), b = 5.0976 (5), c = 27.698 (4) Å and β = 105.729 (6)°. After dynamical refinement, its absolute structure was determined by comparing the R factors and calculating the z-scores of the two possible enanti­omorphs of beauveriolide I.

This report presents a comprehensive investigation into the synthesis and characterization of Schiff base com­pounds derived from benzene­sul­fon­amide. The synthesis process, involved the reaction between N-cyclo­amino-2-sulf­anil­amide and various substituted o-salicyl­aldehydes, resulted in a set of com­pounds that were subjected to rigorous characterization using advanced spectral techniques, including 1H NMR, 13C NMR and FT–IR spectroscopy, and single-crystal X-ray diffraction. Furthermore, an in-depth assessment of the synthesized com­pounds was conducted through Absorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) analysis, in conjunction with docking studies, to elucidate their pharmacokinetic profiles and potential. Impressively, the ADMET analysis showcased encouraging drug-likeness properties of the newly synthesized Schiff bases. These computational findings were substanti­ated by mol­ecular properties derived from density functional theory (DFT) calculations using the B3LYP/6-31G* method within the Jaguar Module of Schrödinger 2023-2 from Maestro (Schrodinger LLC, New York, USA). The ex­plor­ation of frontier mol­ecular orbitals (HOMO and LUMO) enabled the computation of global reactivity descriptors (GRDs), encompassing charge separation (Egap) and global softness (S). Notably, within this analysis, one Schiff base, namely, 4-bromo-2-{N-[2-(pyr­rol­idine-1-sul­fonyl)phenyl]car­box­imid­oyl}phenol, 20, em­erged with the smallest charge separation (ΔEgap = 3.5780 eV), signifying heightened potential for biological properties. Conversely, 4-bromo-2-{N-[2-(piper­idine-1-sul­fonyl)phenyl]car­box­imid­oyl}phenol, 17, exhibited the largest charge separation (ΔEgap = 4.9242 eV), implying a relatively lower propensity for biological activity. Moreover, the synthesized Schiff bases displayed re­marke­able inhibition of tankyrase poly(ADP-ribose) polymerase enzymes, integral in colon cancer, surpassing the efficacy of a standard drug used for the same purpose. Additionally, their bioavailability scores aligned closely with established medications such as trifluridine and 5-fluoro­uracil. The ex­plor­ation of mol­ecular electrostatic potential through colour mapping delved into the electronic behaviour and reactivity tendencies intrinsic to this diverse range of mol­ecules.

X-ray and electron diffraction methods independently identify the S-enanti­omer of Berkecoumarin [systematic name: (S)-8-hy­droxy-3-(2-hy­droxy­prop­yl)-6-meth­oxy-2H-chromen-2-one]. Isolated from Berkeley Pit Lake Penicillium sp., Berkecoumarin is a natural product with a light-atom com­position (C13H14O5) that challenges in-house absolute structure determination by anomalous scattering. This study further demonstrates the utility of dynamical refinement of electron-diffraction data for absolute structure determination.

The synthesis and structural characterization of three families of coordination com­plexes synthesized from 4'-phenyl-2,2':6',2''-terpyridine (8, Ph-TPY), 4'-(4-chloro­phen­yl)-2,2':6',2''-terpyridine (9, ClPh-TPY) and 4'-(4-meth­oxy­phen­yl)-2,2':6',2''-terpyridine (10, MeOPh-TPY) ligands with the divalent metals Co2+, Fe2+, Mn2+ and Ni2+ are reported. The com­pounds were synthesized from a 1:2 mixture of the metal and ligand, resulting in a series of com­plexes with the general formula [M(R-TPY)2](ClO4)2 (where M = Co2+, Fe2+, Mn2+ and Ni2+, and R-TPY = Ph-TPY, ClPh-TPY and MeOPh-TPY). The general formula and structural and supra­molecular features were determinated by single-crystal X-ray diffraction for bis­(4'-phenyl-2,2':6',2''-terpyridine)­nickel(II) bis­(per­chlo­rate), [Ni(C21H15N3)2](ClO4)2 or [Ni(Ph-TPY)2](ClO4)2, bis­[4'-(4-meth­oxy­phen­yl)-2,2':6',2''-terpyridine]­manganese(II) bis­(per­chlo­rate), [Mn(C22H17N3O)2](ClO4)2 or [Mn(MeOPh-TPY)2](ClO4)2, and bis­(4'-phenyl-2,2':6',2''-ter­py­ridine)­manganese(II) bis­(per­chlo­rate), [Mn(C21H15N3)2](ClO4)2 or [Mn(Ph-TPY)2](ClO4)2. In all three cases, the com­plexes present distorted octa­hedral coordination polyhedra and the crystal packing is determined mainly by weak C—H⋯π inter­actions. All the com­pounds (except for the Ni derivatives, for which FT–IR, UV–Vis and thermal analysis are reported) were fully characterized by spectroscopic (FT–IR, UV–Vis and NMR spectroscopy) and thermal (TGA–DSC, thermogravimetric analysis–differential scanning calorimetry) methods.

Starting from [Mn(C5H4Br)(PPh3)(CO)2] (1a), the cymantrenyl thio­ethers [Mn(C5H4SMe)(PPh3)(CO)2] (1b) and [Mn{C5H4–nBr(SMe)n}(PPh3)(CO)2] (n = 1 for com­pound 2, n = 2 for 3 and n = 3 for 4) were obtained, using either n-butyllithium (n-BuLi), lithium diiso­propyl­amide (LDA) or lithium tetra­methyl­piperidide (LiTMP) as base, followed by electrophilic quenching with MeSSMe. Stepwise consecutive reaction of [Mn(C5Br5)(PPh3)(CO)2] with n-BuLi and MeSSMe led finally to [Mn{C5(SMe)5}(PPh3)(CO)2] (11), only the fifth com­plex to be reported containing a perthiol­ated cyclo­penta­dienyl ring. The mol­ecular and crystal structures of 1b, 3, 4 and 11 were determined and were studied for the occurrence of S⋯S and S⋯Br inter­actions. It turned out that although some inter­actions of this type occurred, they were of minor importance for the arrangement of the mol­ecules in the crystal.

The title compound, 3-[(benzo-1,3-dioxol-5-yl)amino]-4-meth­oxy­cyclo­but-3-ene-1,2-dione, C12H9NO5 (3), is a precursor to an anti­mycobacterial squaramide. Block-shaped crystals of a monoclinic form (3-I, space group P21/c, Z = 8, Z' = 2) and needle-shaped crystals of a triclinic form (3-II, space group P-1, Z = 4, Z' = 2) were found to crystallize concomitantly. In both crystal forms, R22(10) dimers assemble through N—H⋯O=C hydrogen bonds. These dimers are formed from crystallographically unique mol­ecules in 3-I, but exhibit crystallographic Ci symmetry in 3-II. Twinning by pseudomerohedry was encountered in the crystals of 3-II. The conformations of 3 in the solid forms 3-I and 3-II are different from one another but are similar for the unique mol­ecules in each polymorph. Density functional theory (DFT) calculations on the free mol­ecule of 3 indicate that a nearly planar conformation is preferred.

Chalcopyrite, the world's primary copper ore mineral, is abundant in Latin America. Copper extraction offers significant economic and social benefits due to its strategic importance across various industries. However, the hydro­metallurgical route, considered more environmentally friendly for processing low-grade chal­co­py­rite ores, remains challenging, as does its concentration by froth flotation. This limited understanding stems from the poorly understood structure and reactivity of chal­co­py­rite surfaces. This study reviews recent contributions using density functional theory (DFT) calculations with periodic boundary conditions and slab models to elucidate chal­co­py­rite surface properties. Our analysis reveals that reconstructed surfaces preferentially expose S atoms at the topmost layer. Furthermore, some studies report the formation of di­sulfide groups (S22−) on pristine sulfur-terminated surfaces, accom­panied by the reduction of Fe3+ to Fe2+, likely due to surface oxidation. Additionally, Fe sites are consistently identified as favourable adsorption locations for both oxygen (O2) and water (H2O) mol­ecules. Finally, the potential of com­puter modelling for investigating collector–chal­co­py­rite surface inter­actions in the context of selective froth flotation is discussed, highlighting the need for further research in this area.

A series of organotin heterocycles of general formula [{Me2C(C6H3CH2)2O}SnR2] [R = methyl (Me, 4), n-butyl (n-Bu, 5), benzyl (Bn, 6) and phenyl (Ph, 7)] was easily synthesized by a Barbier-type reaction assisted by the sonochemical activation of metallic magnesium. The 119Sn{1H} NMR data for all four com­pounds confirm the presence of a central Sn atom in a four-coordinated environment in solution. Single-crystal X-ray diffraction studies for 17,17-dimethyl-7,7-di­phenyl-15-oxa-7-stanna­tetra­cyclo­[11.3.1.05,16.09,14]hepta­deca-1,3,5(16),9(14),10,12-hexa­­ene, [Sn(C6H5)2(C17H16O)], 7, at 100 and 295 K con­firmed the formation of a mono­nuclear eight-membered heterocycle, with a conformation depicted as boat–chair, resulting in a weak Sn⋯O inter­action. The Sn and O atoms are surrounded by hydro­phobic C—H bonds. A Hirshfeld surface analysis of 7 showed that the eight-membered heterocycles are linked by weak C—H⋯π, π–π and H⋯H noncovalent inter­actions. The pairwise inter­action energies showed that the cohesion between the heterocycles are mainly due to dispersion forces.

It is well known that Hirshfeld surfaces provide an easy and straightforward way of analysing inter­molecular inter­actions in the crystal environment. The use of atomic Hirshfeld surfaces has also demonstrated that such surfaces carry information related to chemical bonds which allow a deeper evaluation of the structures. Here we briefly summarize the approach of atomic Hirshfeld surfaces while further evaluating the kind of information that can be retrieved from them. We show that the analysis of the metal-centre Hirshfeld surfaces from structures refined via Hirshfeld Atom Refinement (HAR) allow accurate evaluation of contacts of type M⋯H, and that such contacts can be related to the overall shape of the surfaces. The com­pounds analysed were tetra­aqua­bis­(3-carb­oxy­propionato)metal(II), [M(C4H3O4)2(H2O)4], for metal(II)/M = manganese/Mn, cobalt/Co, nickel/Ni and zinc/Zn. We also evaluate the sensitivity of the surfaces by an investigation of seemingly flat surfaces through analysis of the curvature functions in the direction of C—C bonds. The obtained values not only demonstrate variations in curvature but also show a correlation with the hybridization of the C atoms involved in the bond.

Three 2,4-di­aryl­pyrroles were synthesized starting from 4-nitro­butano­nes and the crystal structures of two derivatives were analysed. These are 4-(4-meth­oxy­phen­yl)-2-(thio­phen-2-yl)-1H-pyrrole, C15H13NOS, and 3-(4-bromo­phen­yl)-2-nitroso-5-phenyl-1H-pyrrole, C16H11BrN2O. Although pyrroles without sub­stituents at the α-position with respect to the N atom are very air sensitive and tend to polymerize, we succeeded in growing an adequate crystal for X-ray diffraction analysis. Further derivatization using sodium nitrite afforded a nitrosyl pyrrole derivative, which crystallized in the triclinic space group Poverline{1} with Z = 6. Thus, herein we report the first crystal structure of a nitrosyl pyrrole. Inter­estingly, the co-operative hydrogen bonds in this NO-substituted pyrrole lead to a trimeric structure with bifurcated halogen bonds at the ends, forming a two-dimensional (2D) layer with inter­stitial voids having a radius of 5 Å, similar to some reported macrocyclic porphyrins.

The crystal structures of two coordination com­pounds, (acetato-κO)(2,2'-bi­py­ri­dine-κ2N,N')(1,10-phenanthroline-κ2N,N')copper(II) acetate hexa­hydrate, [Cu(C2H3O2)(C10H8N2)(C12H8N2)](C2H3O2)·6H2O or [Cu(bipy)(phen)Ac]Ac·6H2O, and (acetato-κO)bis­(2,2'-bi­py­ri­dine-κ2N,N')copper(II) acetate–acetic acid–water (1/1/3), [Cu(C2H3O2)(C10H8N2)2](C2H3O2)·C2H4O2·3H2O or [Cu(bipy)2Ac]Ac·HAc·3H2O, are reported and com­pared with the previously published structure of [Cu(phen)2Ac]Ac·7H2O (phen is 1,10-phenanthroline, bipy for 2,2'-bi­py­ri­dine, ac is acetate and Hac is acetic acid). The geometry around the metal centre is penta­coordinated, but highly distorted in all three cases. The coordination number and the geometric distortion are both discussed in detail, and all com­plexes belong to the space group Poverline{1}. The analysis of the geometric parameters and the Hirshfeld surface properties dnorm and curvedness provide information about the metal–ligand inter­actions in these com­plexes and allow com­parison with similar systems.

In this study, we report the results of continuous rotation electron diffraction studies of single DyPO4·nH2O (rhabdophane) nanocrystals. The diffraction patterns can be fit to a trigonal lattice (P3121) with lattice parameters a = 7.019 (5) and c = 6.417 (5) Å. However, there is also a set of diffuse background scattering features present that are associated with a disordered superstructure that is double these lattice parameters and fits with an arrangement of water mol­ecules present in the structure pore. Pair distribution function (PDF) maps based on the diffuse background allowed the extent of the water correlation to be estimated, with 2–3 nm correlation along the c axis and ∼5 nm along the a/b axis.

Catalase is an antioxidant enzyme that breaks down hydrogen peroxide (H2O2) into molecular oxygen and water. In all monofunctional catalases the pathway that H2O2 takes to the catalytic centre is via the `main channel'. However, the structure of this channel differs in large-subunit and small-subunit catalases. In large-subunit catalases the channel is 15 Å longer and consists of two distinct parts, including a hydrophobic lower region near the heme and a hydrophilic upper region where multiple H2O2 routes are possible. Conserved glutamic acid and threonine residues are located near the intersection of these two regions. Mutations of these two residues in the Scytalidium thermophilum catalase had no significant effect on catalase activity. However, the secondary phenol oxidase activity was markedly altered, with kcat and kcat/Km values that were significantly increased in the five variants E484A, E484I, T188D, T188I and T188F. These variants also showed a lower affinity for inhibitors of oxidase activity than the wild-type enzyme and a higher affinity for phenolic substrates. Oxidation of heme b to heme d did not occur in most of the studied variants. Structural changes in solvent-chain integrity and channel architecture were also observed. In summary, modification of the main-channel gate glutamic acid and threonine residues has a greater influence on the secondary activity of the catalase enzyme, and the oxidation of heme b to heme d is predominantly inhibited by their conversion to aliphatic and aromatic residues.

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 predicted structural models of sufficient accuracy provide a powerful approach to molecular replacement. Taking advantage of these advances, AlphaFold predictions were applied to enable structure determination of a bacterial protein of unknown function (UniProtKB Q63NT7, NCBI locus BPSS0212) based on diffraction data that had evaded phasing attempts using MIR and anomalous scattering methods. Using both X-ray and micro-electron (microED) diffraction data, it was possible to solve the structure of the main fragment of the protein using a predicted model of that domain as a starting point. The use of predicted structural models importantly expands the promise of electron diffraction, where structure determination relies critically on molecular replacement.

Low-molecular-weight (LMW) thiols are involved in many processes in all organisms, playing a protective role against reactive species, heavy metals, toxins and antibiotics. Actinobacteria, such as Mycobacterium tuberculosis, use the LMW thiol mycothiol (MSH) to buffer the intracellular redox environment. The NADPH-dependent FAD-containing oxidoreductase mycothiol disulfide reductase (Mtr) is known to reduce oxidized mycothiol disulfide (MSSM) to MSH, which is crucial to maintain the cellular redox balance. In this work, the first crystal structures of Mtr are presented, expanding the structural knowledge and understanding of LMW thiol reductases. The structural analyses and docking calculations provide insight into the nature of Mtrs, with regard to the binding and reduction of the MSSM substrate, in the context of related oxidoreductases. The putative binding site for MSSM suggests a similar binding to that described for the homologous glutathione reductase and its respective substrate glutathione disulfide, but with distinct structural differences shaped to fit the bulkier MSSM substrate, assigning Mtrs as uniquely functioning reductases. As MSH has been acknowledged as an attractive antitubercular target, the structural findings presented in this work may contribute towards future antituberculosis drug development.

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 adds to the understanding of the structural basis of enzyme inhibition and thermostability in mevalonate kinases.

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 crystallography at 1.78 and 1.61 Å resolution, respectively. It is also shown that despite coordinating a single non-natural metal cation, the PTE-R18 mutant is still able to maintain esterase activity.

The formation of a vitrified thin film embedded with randomly oriented macromolecules is an essential prerequisite for cryogenic sample electron microscopy. Most commonly, this is achieved using the plunge-freeze method first described nearly 40 years ago. Although this is a robust method, the behaviour of different macromolecules shows great variation upon freezing and often needs to be optimized to obtain an isotropic, high-resolution reconstruction. For a macromolecule in such a film, the probability of encountering the air–water interface in the time between blotting and freezing and adopting preferred orientations is very high. 3D reconstruction using preferentially oriented particles often leads to anisotropic and uninterpretable maps. Currently, there are no general solutions to this prevalent issue, but several approaches largely focusing on sample preparation with the use of additives and novel grid modifications have been attempted. In this study, the effect of physical and chemical factors on the orientations of macromolecules was investigated through an analysis of selected well studied macromolecules, and important parameters that determine the behaviour of proteins on cryo-EM grids were revealed. These insights highlight the nature of the interactions that cause preferred orientations and can be utilized to systematically address orientation bias for any given macromolecule and to provide a framework to design small-molecule additives to enhance sample stability and behaviour.

The Azotobacter vinelandii FeSII protein forms an oxygen-resistant complex with the nitrogenase MoFe and Fe proteins. FeSII is an adrenodoxin-type ferredoxin that forms a dimer in solution. Previously, the crystal structure was solved [Schlesier et al. (2016), J. Am. Chem. Soc. 138, 239–247] with five copies in the asymmetric unit. One copy is a normal adrenodoxin domain that forms a dimer with its crystallographic symmetry mate. The other four copies are in an `open' conformation with a loop flipped out exposing the 2Fe–2S cluster. The open and closed conformations were interpreted as oxidized and reduced, respectively, and the large conformational change in the open configuration allowed binding to nitrogenase. Here, the structure of FeSII was independently solved in the same crystal form. The positioning of the atoms in the unit cell is similar to the earlier report. However, the interpretation of the structure is different. The `open' conformation is interpreted as the product of a crystallization-induced domain swap. The 2Fe–2S cluster is not exposed to solvent, but in the crystal its interacting helix is replaced by the same helix residues from a crystal symmetry mate. The domain swap is complicated, as it is unusual in being in the middle of the protein rather than at a terminus, and it creates arrangements of molecules that can be interpreted in multiple ways. It is also cautioned that crystal structures should be interpreted in terms of the contents of the entire crystal rather than of one asymmetric unit.

The Mycobacterium tuberculosis trifunctional enzyme (MtTFE) is an α2β2 tetrameric enzyme in which the α-chain harbors the 2E-enoyl-CoA hydratase (ECH) and 3S-hydroxyacyl-CoA dehydrogenase (HAD) active sites, and the β-chain provides the 3-ketoacyl-CoA thiolase (KAT) active site. Linear, medium-chain and long-chain 2E-enoyl-CoA molecules are the preferred substrates of MtTFE. Previous crystallographic binding and modeling studies identified binding sites for the acyl-CoA substrates at the three active sites, as well as the NAD binding pocket at the HAD active site. These studies also identified three additional CoA binding sites on the surface of MtTFE that are different from the active sites. It has been proposed that one of these additional sites could be of functional relevance for the substrate channeling (by surface crawling) of reaction intermediates between the three active sites. Here, 226 fragments were screened in a crystallographic fragment-binding study of MtTFE crystals, resulting in the structures of 16 MtTFE–fragment complexes. Analysis of the 121 fragment-binding events shows that the ECH active site is the `binding hotspot' for the tested fragments, with 41 binding events. The mode of binding of the fragments bound at the active sites provides additional insight into how the long-chain acyl moiety of the substrates can be accommodated at their proposed binding pockets. In addition, the 20 fragment-binding events between the active sites identify potential transient binding sites of reaction intermediates relevant to the possible channeling of substrates between these active sites. These results provide a basis for further studies to understand the functional relevance of the latter binding sites and to identify substrates for which channeling is crucial.

The interpretation of cryo-EM maps often includes the docking of known or predicted structures of the components, which is particularly useful when the map resolution is worse than 4 Å. Although it can be effective to search the entire map to find the best placement of a component, the process can be slow when the maps are large. However, frequently there is a well-founded hypothesis about where particular components are located. In such cases, a local search using a map subvolume will be much faster because the search volume is smaller, and more sensitive because optimizing the search volume for the rotation-search step enhances the signal to noise. A Fourier-space likelihood-based local search approach, based on the previously published em_placement software, has been implemented in the new emplace_local program. Tests confirm that the local search approach enhances the speed and sensitivity of the computations. An interactive graphical interface in the ChimeraX molecular-graphics program provides a convenient way to set up and evaluate docking calculations, particularly in defining the part of the map into which the components should be placed.

Eukaryotic TIR (Toll/interleukin-1 receptor protein) domains signal via TIR–TIR interactions, either by self-association or by interaction with other TIR domains. In mammals, TIR domains are found in Toll-like receptors (TLRs) and cytoplasmic adaptor proteins involved in pro-inflammatory signaling. Previous work revealed that the MAL TIR domain (MALTIR) nucleates the assembly of MyD88TIR into crystalline arrays in vitro. A microcrystal electron diffraction (MicroED) structure of the MyD88TIR assembly has previously been solved, revealing a two-stranded higher-order assembly of TIR domains. In this work, it is demonstrated that the TIR domain of TLR2, which is reported to signal as a heterodimer with either TLR1 or TLR6, induces the formation of crystalline higher-order assemblies of MyD88TIR in vitro, whereas TLR1TIR and TLR6TIR do not. Using an improved data-collection protocol, the MicroED structure of TLR2TIR-induced MyD88TIR microcrystals was determined at a higher resolution (2.85 Å) and with higher completeness (89%) compared with the previous structure of the MALTIR-induced MyD88TIR assembly. Both assemblies exhibit conformational differences in several areas that are important for signaling (for example the BB loop and CD loop) compared with their monomeric structures. These data suggest that TLR2TIR and MALTIR interact with MyD88 in an analogous manner during signaling, nucleating MyD88TIR assemblies uni­directionally.

β-Glucosidase from the thermophilic bacterium Caldicellulosiruptor saccharo­lyticus (Bgl1) has been denoted as having an attractive catalytic profile for various industrial applications. Bgl1 catalyses the final step of in the decomposition of cellulose, an unbranched glucose polymer that has attracted the attention of researchers in recent years as it is the most abundant renewable source of reduced carbon in the biosphere. With the aim of enhancing the thermostability of Bgl1 for a broad spectrum of biotechnological processes, it has been subjected to structural studies. Crystal structures of Bgl1 and its complex with glucose were determined at 1.47 and 1.95 Å resolution, respectively. Bgl1 is a member of glycosyl hydrolase family 1 (GH1 superfamily, EC 3.2.1.21) and the results showed that the 3D structure of Bgl1 follows the overall architecture of the GH1 family, with a classical (β/α)8 TIM-barrel fold. Comparisons of Bgl1 with sequence or structural homologues of β-glucosidase reveal quite similar structures but also unique structural features in Bgl1 with plausible functional roles.

Our study compares short-range order parameters refined from the diffuse scattering in single-crystal X-ray and single-crystal electron diffraction data. Nb0.84CoSb was chosen as a reference material. The correlations between neighbouring vacancies and the displacements of Sb and Co atoms were refined from the diffuse scattering using a Monte Carlo refinement in DISCUS. The difference between the Sb and Co displacements refined from the diffuse scattering and the Sb and Co displacements refined from the Bragg reflections in single-crystal X-ray diffraction data is 0.012 (7) Å for the refinement on diffuse scattering in single-crystal X-ray diffraction data and 0.03 (2) Å for the refinement on the diffuse scattering in single-crystal electron diffraction data. As electron diffraction requires much smaller crystals than X-ray diffraction, this opens up the possibility of refining short-range order parameters in many technologically relevant materials for which no crystals large enough for single-crystal X-ray diffraction are available.

Molecular structures can be determined in vitro and in situ with cryo-electron microscopy (cryo-EM). Specimen preparation is a major obstacle in cryo-EM. Typical sample preparation is orders of magnitude slower than biological processes. Time-resolved cryo-EM (TR-cryo-EM) can capture short-lived states. Here, Cryo-EM sample preparation with light-activated molecules (C-SPAM) is presented, an open-source, photochemistry-coupled device for TR-cryo-EM that enables millisecond resolution and tunable timescales across broad biological applications.

As an important characterization method, pair distribution function (PDF) has been extensively used in structural analysis of nanomaterials, providing key insights into the degree of crystallinity, atomic structure, local disorder etc. The collection of scattering signals with good statistics is necessary for a reliable structural analysis. However, current conventional electron diffraction experiments using PDF (ePDF) are limited in their ability to acquire continuous diffraction rings for large nanoparticles. Herein, a new method – tilt-ePDF – is proposed to improve the data quality and compatibility of ePDF by a combination of electron diffraction and specimen tilting. In the present work, a tilt-series of electron diffraction patterns was collected from gold nanoparticles with three different sizes and a standard sample polycrystalline aluminium film for ePDF analysis. The results show that tilt-ePDF can not only enhance the continuity of diffraction rings, but can also improve the signal-to-noise ratio in the high scattering angle range. As a result, compared with conventional ePDF data, tilt-ePDF data provide structure parameters with a better accuracy and lower residual factors in the refinement against the crystal structure. This method provides a new way of utilizing ePDF to obtain accurate local structure information from nanoparticles.

The discovery of lytic polysaccharide monooxygenases (LPMOs), a family of copper-dependent enzymes that play a major role in polysaccharide degradation, has revealed the importance of oxidoreductases in the biological utilization of biomass. In fungi, a range of redox proteins have been implicated as working in harness with LPMOs to bring about polysaccharide oxidation. In bacteria, less is known about the interplay between redox proteins and LPMOs, or how the interaction between the two contributes to polysaccharide degradation. We therefore set out to characterize two previously unstudied proteins from the shipworm symbiont Teredinibacter turnerae that were initially identified by the presence of carbohydrate binding domains appended to uncharacterized domains with probable redox functions. Here, X-ray crystal structures of several domains from these proteins are presented together with initial efforts to characterize their functions. The analysis suggests that the target proteins are unlikely to function as LPMO electron donors, raising new questions as to the potential redox functions that these large extracellular multi-haem-containing c-type cytochromes may perform in these bacteria.

Form factors based on aspherical models of atomic electron density have brought great improvement in the accuracies of hydrogen atom parameters derived from X-ray crystal structure refinement. Today, two main groups of such models are available, the banks of transferable atomic densities parametrized using the Hansen–Coppens multipole model which allows for rapid evaluation of atomic form factors and Hirshfeld atom refinement (HAR)-related methods which are usually more accurate but also slower. In this work, a model that combines the ideas utilized in the two approaches is tested. It uses atomic electron densities based on Hirshfeld partitions of electron densities, which are precalculated and stored in a databank. This model was also applied during the refinement of the structures of five small molecules. A comparison of the resulting hydrogen atom parameters with those derived from neutron diffraction data indicates that they are more accurate than those obtained with the Hansen–Coppens based databank, and only slightly less accurate than those obtained with a version of HAR that neglects the crystal environment. The advantage of using HAR becomes more noticeable when the effects of the environment are included. To speed up calculations, atomic densities were represented by multipole expansion with spherical harmonics up to l = 7, which used numerical radial functions (a different approach to that applied in the Hansen–Coppens model). Calculations of atomic form factors for the small protein crambin (at 0.73 Å resolution) took only 68 s using 12 CPU cores.

Dynamical refinement is a well established method for refining crystal structures against 3D electron diffraction (ED) data and its benefits have been discussed in the literature [Palatinus, Petříček & Corrêa, (2015). Acta Cryst. A71, 235–244; Palatinus, Corrêa et al. (2015). Acta Cryst. B71, 740–751]. However, until now, dynamical refinements have only been conducted using the independent atom model (IAM). Recent research has shown that a more accurate description can be achieved by applying the transferable aspherical atom model (TAAM), but this has been limited only to kinematical refinements [Gruza et al. (2020). Acta Cryst. A76, 92–109; Jha et al. (2021). J. Appl. Cryst. 54, 1234–1243]. In this study, we combine dynamical refinement with TAAM for the crystal structure of 1-methyl­uracil, using data from precession ED. Our results show that this approach improves the residual Fourier electrostatic potential and refinement figures of merit. Furthermore, it leads to systematic changes in the atomic displacement parameters of all atoms and the positions of hydrogen atoms. We found that the refinement results are sensitive to the parameters used in the TAAM modelling process. Though our results show that TAAM offers superior performance compared with IAM in all cases, they also show that TAAM parameters obtained by periodic DFT calculations on the refined structure are superior to the TAAM parameters from the UBDB/MATTS database. It appears that multipolar parameters transferred from the database may not be sufficiently accurate to provide a satisfactory description of all details of the electrostatic potential probed by the 3D ED experiment.

Human peptidylarginine deiminase isoform VI (PAD6), which is predominantly limited to cytoplasmic lattices in the mammalian oocytes in ovarian tissue, is essential for female fertility. It belongs to the peptidylarginine deiminase (PAD) enzyme family that catalyzes the conversion of arginine residues to citrulline in proteins. In contrast to other members of the family, recombinant PAD6 was previously found to be catalytically inactive. We sought to provide structural insight into the human homologue to shed light on this observation. We report here the first crystal structure of PAD6, determined at 1.7 Å resolution. PAD6 follows the same domain organization as other structurally known PAD isoenzymes. Further structural analysis and size-exclusion chromatography show that PAD6 behaves as a homodimer similar to PAD4. Differential scanning fluorimetry suggests that PAD6 does not coordinate Ca2+ which agrees with acidic residues found to coordinate Ca2+ in other PAD homologs not being conserved in PAD6. The crystal structure of PAD6 shows similarities with the inactive state of apo PAD2, in which the active site conformation is unsuitable for catalytic citrullination. The putative active site of PAD6 adopts a non-productive conformation that would not allow protein–substrate binding due to steric hindrance with rigid secondary structure elements. This observation is further supported by the lack of activity on the histone H3 and cytokeratin 5 substrates. These findings suggest a different mechanism for enzymatic activation compared with other PADs; alternatively, PAD6 may exert a non-enzymatic function in the cytoplasmic lattice of oocytes and early embryos.

Although COF-300 is often used as an example to study the synthesis and structure of (3D) covalent organic frameworks (COFs), knowledge of the underlying synthetic processes is still fragmented. Here, an optimized synthetic procedure based on a combination of linker protection and modulation was applied. Using this approach, the influence of time and temperature on the synthesis of COF-300 was studied. Synthesis times that were too short produced materials with limited crystallinity and porosity, lacking the typical pore flexibility associated with COF-300. On the other hand, synthesis times that were too long could be characterized by loss of crystallinity and pore order by degradation of the tetrakis(4-aminophenyl)methane (TAM) linker used. The presence of the degradation product was confirmed by visual inspection, Raman spectroscopy and X-ray photoelectron spectroscopy (XPS). As TAM is by far the most popular linker for the synthesis of 3D COFs, this degradation process might be one of the reasons why the development of 3D COFs is still lagging compared with 2D COFs. However, COF crystals obtained via an optimized procedure could be structurally probed using 3D electron diffraction (3DED). The 3DED analysis resulted in a full structure determination of COF-300 at atomic resolution with satisfying data parameters. Comparison of our 3DED-derived structural model with previously reported single-crystal X-ray diffraction data for this material, as well as parameters derived from the Cambridge Structural Database, demonstrates the high accuracy of the 3DED method for structure determination. This validation might accelerate the exploitation of 3DED as a structure determination technique for COFs and other porous materials.

The success of experimental phasing in macromolecular crystallography relies primarily on the accurate locations of heavy atoms bound to the target crystal. To improve the process of substructure determination, a modified phase-retrieval algorithm built on the framework of the relaxed alternating averaged reflection (RAAR) algorithm has been developed. Importantly, the proposed algorithm features a combination of the π-half phase perturbation for weak reflections and enforces the direct-method-based tangent formula for strong reflections in reciprocal space. The proposed algorithm is extensively demonstrated on a total of 100 single-wavelength anomalous diffraction (SAD) experimental datasets, comprising both protein and nucleic acid structures of different qualities. Compared with the standard RAAR algorithm, the modified phase-retrieval algorithm exhibits significantly improved effectiveness and accuracy in SAD substructure determination, highlighting the importance of additional constraints for algorithmic performance. Furthermore, the proposed algorithm can be performed without human intervention under most conditions owing to the self-adaptive property of the input parameters, thus making it convenient to be integrated into the structural determination pipeline. In conjunction with the IPCAS software suite, we demonstrated experimentally that automatic de novo structure determination is possible on the basis of our proposed algorithm.

Spectroscopic data, particularly diffraction data, are essential for materials characterization due to their comprehensive crystallographic information. The current crystallographic phase identification, however, is very time consuming. To address this challenge, we have developed a real-time crystallographic phase identifier based on a convolutional self-attention neural network (CPICANN). Trained on 692 190 simulated powder X-ray diffraction (XRD) patterns from 23 073 distinct inorganic crystallographic information files, CPICANN demonstrates superior phase-identification power. Single-phase identification on simulated XRD patterns yields 98.5 and 87.5% accuracies with and without elemental information, respectively, outperforming JADE software (68.2 and 38.7%, respectively). Bi-phase identification on simulated XRD patterns achieves 84.2 and 51.5% accuracies, respectively. In experimental settings, CPICANN achieves an 80% identification accuracy, surpassing JADE software (61%). Integration of CPICANN into XRD refinement software will significantly advance the cutting-edge technology in XRD materials characterization.

Light–oxygen–voltage (LOV) domains are small photosensory flavoprotein modules that allow the conversion of external stimuli (sunlight) into intra­cellular signals responsible for various cell behaviors (e.g. phototropism and chloro­plast relocation). This ability relies on the light-induced formation of a covalent thio­ether adduct between a flavin chromophore and a reactive cysteine from the protein environment, which triggers a cascade of structural changes that result in the activation of a serine/threonine (Ser/Thr) kinase. Recent developments in time-resolved crystallography may allow the activation cascade of the LOV domain to be observed in real time, which has been elusive. In this study, we report a robust protocol for the production and stable delivery of microcrystals of the LOV domain of phototropin Phot-1 from Chlamydomonas reinhardtii (CrPhotLOV1) with a high-viscosity injector for time-resolved serial synchrotron crystallography (TR-SSX). The detailed process covers all aspects, from sample optimization to data collection, which may serve as a guide for soluble protein preparation for TR-SSX. In addition, we show that the crystals obtained preserve the photoreactivity using infrared spectroscopy. Furthermore, the results of the TR-SSX experiment provide high-resolution insights into structural alterations of CrPhotLOV1 from Δt = 2.5 ms up to Δt = 95 ms post-photoactivation, including resolving the geometry of the thio­ether adduct and the C-terminal region implicated in the signal transduction process.

This study examines various methods for modelling the electron density and, thus, the electrostatic potential of an organometallic complex for use in crystal structure refinement against 3D electron diffraction (ED) data. It focuses on modelling the scattering factors of iron(III), considering the electron density distribution specific for coordination with organic linkers. We refined the structural model of the metal–organic complex, iron(III) acetyl­acetonate (FeAcAc), using both the independent atom model (IAM) and the transferable aspherical atom model (TAAM). TAAM refinement initially employed multipolar parameters from the MATTS databank for acetyl­acetonate, while iron was modelled with a spherical and neutral approach (TAAM ligand). Later, custom-made TAAM scattering factors for Fe—O coordination were derived from DFT calculations [TAAM-ligand-Fe(III)]. Our findings show that, in this compound, the TAAM scattering factor corresponding to Fe3+ has a lower scattering amplitude than the Fe3+ charged scattering factor described by IAM. When using scattering factors corresponding to the oxidation state of iron, IAM inaccurately represents electrostatic potential maps and overestimates the scattering potential of the iron. In addition, TAAM significantly improved the fitting of the model to the data, shown by improved R1 values, goodness-of-fit (GooF) and reduced noise in the Fourier difference map (based on the residual distribution analysis). For 3D ED, R1 values improved from 19.36% (IAM) to 17.44% (TAAM-ligand) and 17.49% (TAAM-ligand-Fe3+), and for single-crystal X-ray diffraction (SCXRD) from 3.82 to 2.03% and 1.98%, respectively. For 3D ED, the most significant R1 reductions occurred in the low-resolution region (8.65–2.00 Å), dropping from 20.19% (IAM) to 14.67% and 14.89% for TAAM-ligand and TAAM-ligand-Fe(III), respectively, with less improvement in high-resolution ranges (2.00–0.85 Å). This indicates that the major enhancements are due to better scattering modelling in low-resolution zones. Furthermore, when using TAAM instead of IAM, there was a noticeable improvement in the shape of the thermal ellipsoids, which more closely resembled those of an SCXRD-refined model. This study demonstrates the applicability of more sophisticated scattering factors to improve the refinement of metal–organic complexes against 3D ED data, suggesting the need for more accurate modelling methods and highlighting the potential of TAAM in examining the charge distribution of large molecular structures using 3D ED.

Reaching beyond the commonly used spherical atomic electron density model allows one to greatly improve the accuracy of hydrogen atom structural param­eters derived from X-ray data. However, the effects of atomic asphericity are less explored for electron diffraction data. In this work, Hirshfeld atom refinement (HAR), a method that uses an accurate description of electron density by quantum mechanical calculation for a system of interest, was applied for the first time to the kinematical refinement of electron diffraction data. This approach was applied here to derive the structure of ordinary hexagonal ice (Ih). The effect of introducing HAR is much less noticeable than in the case of X-ray refinement and it is largely overshadowed by dynamical scattering effects. It led to only a slight change in the O—H bond lengths (shortening by 0.01 Å) compared with the independent atom model (IAM). The average absolute differences in O—H bond lengths between the kinematical refinements and the reference neutron structure were much larger: 0.044 for IAM and 0.046 Å for HAR. The refinement results changed considerably when dynamical scattering effects were modelled – with extinction correction or with dynamical refinement. The latter led to an improvement of the O—H bond length accuracy to 0.021 Å on average (with IAM refinement). Though there is a potential for deriving more accurate structures using HAR for electron diffraction, modelling of dynamical scattering effects seems to be a necessary step to achieve this. However, at present there is no software to support both HAR and dynamical refinement.

A pressure-induced triclinic-to-monoclinic phase transition has been caught `in the act' over a wider series of high-pressure synchrotron diffraction experiments conducted on a large, photoluminescent organo-gold(I) compound. Here, we describe the mechanism of this single-crystal-to-single-crystal phase transition, the onset of which occurs at ∼0.6 GPa, and we report a high-quality structure of the new monoclinic phase, refined using aspherical atomic scattering factors. Our case illustrates how conducting a fast series of diffraction experiments, enabled by modern equipment at synchrotron facilities, can lead to overestimation of the actual pressure of a phase transition due to slow transformation kinetics.

The TIR (Toll/interleukin-1 receptor) domain represents a vital structural element shared by proteins with roles in immunity signalling pathways across phyla (from humans and plants to bacteria). Decades of research have finally led to identifying the key features of the molecular basis of signalling by these domains, including the formation of open-ended (filamentous) assemblies (responsible for the signalling by cooperative assembly formation mechanism, SCAF) and enzymatic activities involving the cleavage of nucleotides. We present a historical perspective of the research that led to this understanding, highlighting the roles that different structural methods played in this process: X-ray crystallography (including serial crystallography), microED (micro-crystal electron diffraction), NMR (nuclear magnetic resonance) spectroscopy and cryo-EM (cryogenic electron microscopy) involving helical reconstruction and single-particle analysis. This perspective emphasizes the complementarity of different structural approaches.

3D electron diffraction (3DED) is increasingly employed to determine molec­ular and crystal structures from micro-crystals. Indomethacin is a well known, marketed, small-molecule non-steroidal anti-inflammatory drug with eight known polymorphic forms, of which four structures have been elucidated to date. Using 3DED, we determined the structure of a new ninth polymorph, σ, found within an amorphous solid dispersion, a product formulation sometimes used for active pharmaceutical ingredients with poor aqueous solubility. Subsequently, we found that σ indomethacin can be produced from direct solvent evaporation using di­chloro­methane. These results demonstrate the relevance of 3DED within drug development to directly probe product formulations.

MltG, a membrane-bound lytic transglycosyl­ase, has roles in terminating glycan polymerization in peptidoglycan and incorporating glycan chains into the cell wall, making it significant in bacterial cell-wall biosynthesis and remodeling. This study provides the first reported MltG structure from Mycobacterium abscessus (maMltG), a superbug that has high antibiotic resistance. Our structural and biochemical analyses revealed that MltG has a flexible peptidoglycan-binding domain and exists as a monomer in solution. Further, the putative active site of maMltG was disclosed using structural analysis and sequence comparison. Overall, this study contributes to our understanding of the transglycosyl­ation reaction of the MltG family, aiding the design of next-generation antibiotics targeting M. abscessus.

Regolith draws intensive research attention because of its importance as the basis for fabricating materials for future human space exploration. Martian regolith is predicted to consist of defect-rich crystal structures due to long-term space weathering. The present report focuses on the structural differences between defect-rich and defect-poor forsterite (Mg2SiO4) – one of the major phases in Martian regolith. In this work, forsterites were synthesized using reverse strike co-precipitation and high-energy ball milling (BM). Subsequent post-processing was also carried out using BM to enhance the defects. The crystal structures of the samples were characterized by X-ray powder diffraction and total scattering using Cu and synchrotron radiation followed by Rietveld refinement and pair distribution function (PDF) analysis, respectively. The structural models were deduced by density functional theory assisted PDF refinements, describing both long-range and short-range order caused by defects. The Raman spectral features of the synthetic forsterites complement the ab initio simulation for an in-depth understanding of the associated structural defects.

OaPAC is a recently discovered blue-light-using flavin adenosine dinucleotide (BLUF) photoactivated adenylate cyclase from the cyanobacterium Oscillatoria acuminata that uses adenosine triphosphate and translates the light signal into the production of cyclic adenosine monophosphate. Here, we report crystal structures of the enzyme in the absence of its natural substrate determined from room-temperature serial crystallography data collected at both an X-ray free-electron laser and a synchrotron, and we compare these structures with cryo-macromolecular crystallography structures obtained at a synchrotron by us and others. These results reveal slight differences in the structure of the enzyme due to data collection at different temperatures and X-ray sources. We further investigate the effect of the Y6W mutation in the BLUF domain, a mutation which results in a rearrangement of the hydrogen-bond network around the flavin and a notable rotation of the side chain of the critical Gln48 residue. These studies pave the way for picosecond–millisecond time-resolved serial crystallography experiments at X-ray free-electron lasers and synchrotrons in order to determine the early structural intermediates and correlate them with the well studied pico­second–millisecond spectroscopic intermediates.

Single-crystal X-ray diffraction analysis of small molecule active pharmaceutical ingredients is a key technique in the confirmation of molecular connectivity, including absolute stereochemistry, as well as the solid-state form. However, accessing single crystals suitable for X-ray diffraction analysis of an active pharmaceutical ingredient can be experimentally laborious, especially considering the potential for multiple solid-state forms (solvates, hydrates and polymorphs). In recent years, methods for the exploration of experimental crystallization space of small molecules have undergone a `step-change', resulting in new high-throughput techniques becoming available. Here, the application of high-throughput encapsulated nanodroplet crystallization to a series of six di­hydro­pyridines, calcium channel blockers used in the treatment of hypertension related diseases, is described. This approach allowed 288 individual crystallization experiments to be performed in parallel on each molecule, resulting in rapid access to crystals and subsequent crystal structures for all six di­hydro­pyridines, as well as revealing a new solvate polymorph of nifedipine (1,4-dioxane solvate) and the first known solvate of nimodipine (DMSO solvate). This work further demonstrates the power of modern high-throughput crystallization methods in the exploration of the solid-state landscape of active pharmaceutical ingredients to facilitate crystal form discovery and structural analysis by single-crystal X-ray diffraction.

Traditional X-ray methods are extensively applied to commercial cement samples in order to determine their physical and chemical properties. Powder patterns are routinely used to quantify the composition of these phase mixtures, but structure determination becomes difficult because of reflection overlapping caused by the high number of different crystal structures. The fast-growing 3D electron diffraction technique and its related automated acquisition protocols arise as a potentially very interesting tool for the cement industry, since they enable the fast and systematic acquisition of diffraction data from individual particles. In this context, electron diffraction has been used in the investigation of the different crystalline phases present in various commercial clinkers for cement. Automated data collection procedures and subsequent data processing have enabled the structural characterization of the different crystal structures from which the α'H polymorph of Ca2SiO4 (belite) exhibited satellite reflections. Its average crystal structure has been known since 1971 and satellite reflections have been reported previously, yet the modulation was never fully described by means of the superspace formalism. Here, the incommensurately modulated structure is solved and refined using harmonic and crenel functions in the superspace group Pnma(α00)0ss, showing the potential of 3D electron diffraction for systematic crystallographic characterizations of cement. A full description of the different belite polymorphs is provided considering this modulated structure.

The compression behavior of [Rb(18-crown-6)][SbCl6] crystal under pressure up to 2.16 (3) GPa was investigated in a diamond anvil cell (DAC) using a mixture of pentane–iso­pentane (1:4) as the pressure-transmitting fluid. The compound crystallizes in trigonal space group R3 and no phase transition was observed in the indicated pressure range. The low value of pressure bulk modulus [9.1 (5) GPa] found in this crystal is a characteristic of soft materials with predominant dispersive and electrostatic interaction forces. The nonlinear relationship between unit-cell parameters under high pressure is attributed to the influence of reduced intermolecular H⋯Cl contacts under pressure over 0.73 GPa. It also explains the high compression efficiency of [Rb(18-crown-6)][SbCl6] crystals at relatively low pressures, resulting in a significant shift of the Rb atom to the center of the crown ether cavity. At pressures above 0.9 GPa, steric repulsion forces begin to play a remarkable role, since an increasing number of interatomic H⋯Cl and H⋯H contacts become shorter than the sum of their van der Waals (vdW) radii. Below 0.9 GPa, both unit-cell parameter dependences (P–a and P–c) exhibit hysteresis upon pressure release, demonstrating their influence on the disordered model of Rb atoms. The void reduction under pressure also demonstrates two linear sections with the inflection point at 0.9 GPa. Compression of the crystal is accompanied by a significant decrease in the volume of the voids, leading to the rapid approach of Rb atoms to the center of the crown ether cavity. For the Rb atom to penetrate into the center of the crown ether cavity in [Rb(18-crown-6)][SbCl6], it is necessary to apply a pressure of about 2.5 GPa to disrupt the balance of atomic forces in the crystal. This sample serves as a compression model demonstrating the influence of both attractive and repulsive forces on the change in unit-cell parameters under pressure.