Crystal Contacts Research Articles

How Do Gepotidacin and Zoliflodacin Stabilize DNA Cleavage Complexes with Bacterial Type IIA Topoisomerases? 1. Experimental Definition of Metal Binding Sites

One of the challenges for experimental structural biology in the 21st century is to see chemical reactions happen. Staphylococcus aureus (S. aureus) DNA gyrase is a type IIA topoisomerase that can create temporary double-stranded DNA breaks to regulate DNA topology. Drugs, such as gepotidacin, zoliflodacin and the quinolone moxifloxacin, can stabilize these normally transient DNA strand breaks and kill bacteria. Crystal structures of uncleaved DNA with a gepotidacin precursor (2.1 Å GSK2999423) or with doubly cleaved DNA and zoliflodacin (or with its progenitor QPT-1) have been solved in the same P61 space-group (a = b ≈ 93 Å, c ≈ 412 Å). This suggests that it may be possible to observe the two DNA cleavage steps (and two DNA-religation steps) in this P61 space-group. Here, a 2.58 Å anomalous manganese dataset in this crystal form is solved, and four previous crystal structures (1.98 Å, 2.1 Å, 2.5 Å and 2.65 Å) in this crystal form are re-refined to clarify crystal contacts. The structures clearly suggest a single moving metal mechanism—presented in an accompanying (second) paper. A previously published 2.98 Å structure of a yeast topoisomerase II, which has static disorder around a crystallographic twofold axis, was published as containing two metals at one active site. Re-refined coordinates of this 2.98 Å yeast structure are consistent with other type IIA topoisomerase structures in only having one metal ion at each of the two different active sites.

Comparison of two crystal polymorphs of NowGFP reveals a new conformational state trapped by crystal packing.

Crystal polymorphism serves as a strategy to study the conformational flexibility of proteins. However, the relationship between protein crystal packing and protein conformation often remains elusive. In this study, two distinct crystal forms of a green fluorescent protein variant, NowGFP, are compared: a previously identified monoclinic form (space group C2) and a newly discovered orthorhombic form (space group P212121). Comparative analysis reveals that both crystal forms exhibit nearly identical linear assemblies of NowGFP molecules interconnected through similar crystal contacts. However, a notable difference lies in the stacking of these assemblies: parallel in the monoclinic form and perpendicular in the orthorhombic form. This distinct mode of stacking leads to different crystal contacts and induces structural alteration in one of the two molecules within the asymmetric unit of the orthorhombic crystal form. This new conformational state captured by orthorhombic crystal packing exhibits two unique features: a conformational shift of the β-barrel scaffold and a restriction of pH-dependent shifts of the key residue Lys61, which is crucial for the pH-dependent spectral shift of this protein. These findings demonstrate a clear connection between crystal packing and alternative conformational states of proteins, providing insights into how structural variations influence the function of fluorescent proteins.

Cryo-EM reconstruction of oleate hydratase bound to a phospholipid membrane bilayer

Oleate hydratase (OhyA) is a bacterial peripheral membrane protein that catalyzes FAD-dependent water addition to membrane bilayer-embedded unsaturated fatty acids. The opportunistic pathogen Staphylococcus aureus uses OhyA to counteract the innate immune system and support colonization. Many Gram-positive and Gram-negative bacteria in the microbiome also encode OhyA. OhyA is a dimeric flavoenzyme whose carboxy terminus is identified as the membrane binding domain; however, understanding how OhyA binds to cellular membranes is not complete until the membrane-bound structure has been elucidated. All available OhyA structures depict the solution state of the protein outside its functional environment. Here, we employ liposomes to solve the cryo-electron microscopy structure of the functional unit: the OhyA•membrane complex. The protein maintains its structure upon membrane binding and slightly alters the curvature of the liposome surface. OhyA preferentially associates with 20–30 nm liposomes with multiple copies of OhyA dimers assembling on the liposome surface resulting in the formation of higher-order oligomers. Dimer assembly is cooperative and extends along a formed ridge of the liposome. We also solved an OhyA dimer of dimers structure that recapitulates the intermolecular interactions that stabilize the dimer assembly on the membrane bilayer as well as the crystal contacts in the lattice of the OhyA crystal structure. Our work enables visualization of the molecular trajectory of membrane binding for this important interfacial enzyme.

Generation of a high confidence set of domain-domain interface types to guide protein complex structure predictions by AlphaFold.

While the release of AlphaFold (AF) represented a breakthrough for the prediction of protein complex structures, its sensitivity, especially when using full length protein sequences, still remains limited. Modeling success rates might increase if AF predictions were guided by likely interacting protein fragments. This approach requires available sets of highly confident protein-protein interface types. Computational resources, such as 3did, infer interacting globular domain types from observed contacts in protein structures. Assessing the accuracy of these predicted interface types is difficult because we lack hand-curated reference sets of verified domain-domain interface (DDI) types. To improve protein complex modeling of DDIs by AF, we manually inspected 80 randomly selected DDI types from the 3did resource to generate a first reference set of DDI types. Identified cases of DDI type nonapproval (40%) primarily resulted from inaccurate Pfam domain matches, crystal contacts, and synthetic protein constructs. Using logistic regression, we predicted a subset of 2411 out of 5724 considered DDI types in 3did to be of high confidence, which we subsequently applied to 53000 human-protein interactions to predict DDIs followed by AF modeling. We obtained highly confident AF models for 604 out of 1129 predicted DDIs. Of note, for 47% of them no confident AF structural model could be obtained using full length protein sequences. Code is available at https://github.com/KatjaLuckLab/DDI_manuscript.

Single-point substitution F97M leads to in cellulo crystallization of the biphotochromic protein moxSAASoti

To enhance the photoconversion performance of biphotochromic moxSAASoti protein, a substitution F97 M was introduced. In addition to enhancing the target properties, this substitution also resulted in the crystallization of the recombinant protein within living HeLa cells (also referred to as in cellulo crystallization). The phenomenon of protein crystallization in living cells is not unique, yet the mechanisms and application of in cellulo crystallization remain significant for further research. However, in cellulo crystallization is atypical for fluorescent proteins and detrimental for their biotechnological application. The objective of this study was to elucidate the underlying mechanisms responsible for the crystallization of moxSAASotiF97Min cellulo. For this purpose, the crystal structure of the green form of biphotochromic protein moxSAASotiF97M was determined at high resolution, which surprisingly has a space group, different from those of parent mSAASotiC21N. The analysis provided allowed to propose a mechanism of new crystal contacts formation, which might be a cause of in cellulo protein crystallization.

High-Performance LiNbO3 Domain Wall Memory Devices with Enhanced Selectivity via Optimized Metal-Semiconductor Contact.

Lithium niobate (LiNbO3) single-crystal nanodevices featuring elevated readout domain wall currents exhibit significant potential for integrated circuits in memory computing applications. Nevertheless, challenges stem from suboptimal electrode-LiNbO3 single crystal contact characteristics, which impact the stability of high currents within these devices. In this work, we concentrate on augmenting the domain wall current by refining the fabrication processes of domain wall random access memory (DWRAM). Each LiNbO3 domain wall nanodevice was fabricated using a self-aligned process. Device performance was significantly enhanced by introducing a 10 nm interlayer between the LiNbO3 and Cu electrodes. A comparative analysis of electrical properties was conducted on devices with interlayers made of chromium (Cr) and titanium (Ti), as well as devices without interlayers. After the introduction of the Ti interlayer, the device's coercive voltage demonstrated an 82% reduction, while the current density showed a remarkable 94-fold increase. A 100 nm sized device with the Ti interlayer underwent positive down-negative up pulse testing, demonstrating a writing time of 82 ns at 8 V and an erasing time of 12 μs at -9 V. These operating speeds are significantly faster than those of devices without interlayers. Moreover, the enhanced devices exhibited symmetrical domain switching hysteresis loops with retention times exceeding 106 s. Notably, the coercive voltage (Vc) dispersion remained narrow after more than 1000 switching cycles. At an elevated temperature of 400 K, the device's on/off ratio was maintained at 105. The device's embedded selector demonstrated an ultrahigh selectivity (>106) across various reading voltages. These results underscore the viability of high-density nanoscale integration of ferroelectric domain wall memory.

Combinatorial and algorithmic complexity of crystal structures

Research subject. Numeral indexes describing the complexity of the system of contacts between structural units in crystal structures. Aim. Development of a complexity index for the system of contacts between periodic structural units based on the indices available for those between structural units in island (molecular) structures. Materials and methods. Structural data were selected from the COD, AMCSD, and CSD crystallographic databases. The system of contacts in the structures was analyzed by the Voronoi–Dirichlet polyhedra (VDP) method in the ToposPro software package. Results. The method of topological analysis of the system of contacts in molecular crystals was adapted to all heterodesmic crystal structures and tested on the structures of compounds of several classes. Target complexity indices were developed. Conclusions. Networks of contacts between periodic structural units are low-dimensional. A generalized structural class for such networks can be derived from the original crystal structure data. The algorithmic complexity of heterodesmic structures is subadditive, in contrast to superadditive combinatorial complexity. For the first time, the number of bearing contacts was calculated between periodic structural units, reflecting the algorithmic complexity of the structure at the appropriate level of structural description.

Rock physical characteristics of deep dolomite under complex geological conditions: A case study of 4th Member of Sinian Dengying Formation in the Sichuan Basin, China

The deep-ultra deep carbonate reservoir in China, commonly subjected to modification of multi-stage diagenesis, has extremely high heterogeneity. Conventional rock physics analysis cannot accurately identify the elastic responses of reservoir. Here, the rock physics properties of the dolomite from the 4th Member of the Sinian Dengying Formation are experimentally measured, and the change law of rock physics characteristics is investigated within the framework of the diagenetic processes by the analysis of the elastic and petrologic characteristics, pore structure, and sedimentary environments. The results show that the differentiated diagenesis results in different pore structure characteristics and micro-texture characteristics of the rock. The microbial dolomite of the algal mound-grain beach facies is subjected to the contemporaneous microbial dolomitization and seepage-reflux dolomitization, penecontemporaneous selective dissolution, burial dolomitization, and hydrothermal dolomitization. The resultant crystalline dolomite is found with one main type of the dolomite crystal contact boundaries, and the dissolution pore is extensive development. The siliceous, muddy, and limy dolomite of the inter-beach sea environment mainly experiences the weak capillary concentration dolomitization, intensive mechanical compaction-induced densification, and burial dolomitization. Such crystalline dolomite is observed with four types of contact boundaries, namely the dolomite contact, clay contact, quartz contact, and calcite contact boundaries, and porosity mostly attributed to residual primary inter-granular or crystalline pores. The samples with the same crystal boundary condition have consistent correlations between the compressional- and shear-wave velocities, and between the compressional-wave velocity and the velocity ratio. Additionally, the variation of the acoustic velocity with effective pressure and the intensity of pore-scale fluid-related dispersion are controlled by the differentiation of pore structure types of the samples. The varied effects of soft pores like micro-cracks on the compressional- and shear-wave velocity causes considerable changes in the relationships between the compressional- and shear-wave velocities, compressional-wave velocity and velocity ratio, and porosity and acoustic velocity. This research is an attempt to demonstrate a novel method for investigating the rock physics variation of rock during the geological process, and the obtained findings can provide the rock physics basis for seismic prediction of the characteristics of deep carbonate reservoirs.

ВЛИЯНИЕ ПРИМЕСЕЙ НА ДЕГРАДАЦИЮ СОЕДИНЕНИЙ В СИСТЕМЕ Al – Au

Typical reasons for degradation of micro–welded bimetallic compounds of the aluminum-gold binary system are considered. These types of compounds are often used in the manufacture of semiconductor products (SCPs) and integrated circuits (ICs). Degradation of compounds, revealed during screen tests or during products operating, leads to failures, which dramatically reduces the reliability of electronic equipment (EE). The majority of degradation processes are based on diffusion phenomena, which are mainly determined by temperature, chemical composition and microstructure of metals and alloys. Impurity elements included in the gold coating have a significant effect on the degradation of the wire connections such as crystal contact pads – traverses of package terminals of SCPs. In this regard, it is worth analyzing the effect of various impurities on the diffusion processes and changes in the microstructure of the coating in order to find out the most dangerous chemical elements and determine their role in the degradation of compounds. The paper presents a review of references on the research subject. In the conclusion, the results of analysis in the field of the influence of impurity elements on the degradation of compounds in Al – Au system are summarized, conclusions are drawn about the key role of grain-boundary diffusion on the degradation of micro-welded bimetallic compounds.

Enhancement of strength and water resistance of macro-defect free （MDF） gypsum modified by pregelatinized starch and hydrogen silicone oil

Gypsum, as a building material with renewable and environmentally friendly characteristics, is severely limited due to its poor mechanical properties and water resistance. Here, this paper investigates pregelatinized starch and hydrogen silicon oil modified macro-defect free (MDF) gypsum composite prepared by pressing. The compressive strength, water absorption, and softening coefficients of foamed MDF gypsum composite were subjected to strict testing. The hydration products, microtopography and pore structure was characterized using XRD, FTIR, SEM and LF-NMR. The result shows that the compressive strength of MDF gypsum composite with 5 % pregelatinized starch was 82.4 MPa, a 44.5% increase compared to the dry blank group, whilst water absorption also rose by 7.8%. Additionally, the compressive strength of MDF gypsum composite with 5 % pregelatinized starch and 1% hydrogen silicon oil was 123.9 MPa, 225.2% higher than that of the dry blank group. The water absorption rate decreases to less than 1.2%. Pregelatinized starch and hydrogen silicone oil assist in the accumulation and arrangement of crystals, while also reducing the space between them. The enhancement mechanism of compressive strength and water resistance of MDF gypsum involves pore filling by pregelatinized starch, as well as crystal contact point modification including stronger adhesion by the pregelatinized starch and hydrophobic properties of hydrogen silicon oil.

Analysis of short contacts in crystals of halogenated amino acids: atom–atom interactions vs. energy frameworks

Crystal packing of several halogenated alanine derivatives was investigated using the combined analysis of short atom–atom contacts and energy framework patterns.

X-ray Crystallography Module in MD Simulation Program Amber 2023. Refining the Models of Protein Crystals.

The MD simulation package Amber offers an attractive platform to refine crystallographic structures of proteins: (i) state-of-the-art force fields help to regularize protein coordinates and reconstruct the poorly diffracting elements of the structure, such as flexible loops; (ii) MD simulations restrained by the experimental diffraction data provide an effective strategy to optimize structural models of protein crystals, including explicitly modeled interstitial solvent as well as crystal contacts. Here, we present the new crystallography module xray, released as a part of the Amber 2023 package. This module contains functions to calculate and scale structure factors (including the contributions from bulk solvent), evaluate the maximum-likelihood-type crystallographic potential, and compute its derivative forces. The X-ray functionality of Amber no longer relies on external dependencies so that the full advantage of GPU acceleration can be taken. This makes it possible to refine in a short time hundreds of crystal models, including supercell models comprised of multiple unit cells. The new automated Amber-based refinement procedure leads to an appreciable improvement in Rfree (in some cases, by as much as 0.067) as well as MolProbity scores.

Assembly Requirements for the Construction of Large-Scale Binary Protein Structures.

The precise assembly of multiple biomacromolecules into well-defined structures and materials is of great importance for various biomedical and nanobiotechnological applications. In this study, we investigate the assembly requirements for two-component materials using charged protein nanocages as building blocks. To achieve this, we designed several variants of ferritin nanocages to determine the surface characteristics necessary for the formation of large-scale binary three-dimensional (3D) assemblies. These nanocage variants were employed in protein crystallization experiments and macromolecular crystallography analyses, complemented by computational methods. Through the screening of nanocage variant combinations at various ionic strengths, we identified three essential features for successful assembly: (1) the presence of a favored crystal contact region, (2) the presence of a charged patch not involved in crystal contacts, and (3) sufficient distinctiveness between the nanocages. Surprisingly, the absence of noncrystal contact mediating patches had a detrimental effect on the assemblies, highlighting their unexpected importance. Intriguingly, we observed the formation of not only binary structures but also both negatively and positively charged unitary structures under previously exclusively binary conditions. Overall, our findings will inform future design strategies by providing some design rules, showcasing the utility of supercharging symmetric building blocks in facilitating the assembly of biomacromolecules into large-scale binary 3D assemblies.

Combined NMR and molecular dynamics conformational filter identifies unambiguously dynamic ensembles of Dengue protease NS2B/NS3pro

The dengue protease NS2B/NS3pro has been reported to adopt either an ‘open’ or a ‘closed’ conformation. We have developed a conformational filter that combines NMR with MD simulations to identify conformational ensembles that dominate in solution. Experimental values derived from relaxation parameters for the backbone and methyl side chains were compared with the corresponding back-calculated relaxation parameters of different conformational ensembles obtained from free MD simulations. Our results demonstrate a high prevalence for the ‘closed’ conformational ensemble while the ‘open’ conformation is absent, indicating that the latter conformation is most probably due to crystal contacts. Conversely, conformational ensembles in which the positioning of the co-factor NS2B results in a ‘partially’ open conformation, previously described in both MD simulations and X-ray studies, were identified by our conformational filter. Altogether, we believe that our approach allows for unambiguous identification of true conformational ensembles, an essential step for reliable drug discovery.

Engineering Atomic-Scale Patterning and Resistive Switching in 2D Crystals and Application in Image Processing.

The ultrathin thickness of 2D layered materials affords the control of their properties through defects, surface modification, and electrostatic fields more efficiently compared with bulk architecture. In particular, patterning design, such as moiré superlattice patterns and spatially periodic dielectric structures, are demonstrated to possess the ability to precisely control the local atomic and electronic environment at large scale, thus providing extra degrees of freedom to realize tailored material properties and device functionality. Here,the scalable atomic-scale patterning in superionic cuprous telluride by using the bonding difference at nonequivalent copper sites is reported. Moreover, benefitting from the natural coupling of ordered and disordered sublattices,controllable piezoelectricity-like multilevel switching and bipolar switching with the designed crystal structure and electrical contact is realized, and their application in image enhancement is demonstrated. This work extends the known classes of patternable crystals and atomic switching devices, and ushers in a frontier for image processing with memristors.

INFUENCE OF ADSORPTION LAYERS OF POLYELECTROLYTES ON THE WETTING AND MODIFICATION OF THE POLYSTYRENE SURFACE

The adsorption of cationic and anionic polyelectrolytes from aqueous solutions on the polystyrene surface was studied by quartz crystal microbalance and contact angle measurements. It is shown that, despite the multilayer nature of adsorption, only a monolayer of polyelectrolyte macromolecules is rmly retained on the polymer surface, and the polystyrene surface is hydrophilized. It is shown that with complete monolayer lling of the surface, the degree of modi cation of the polystyrene surface does not depend on the nature of the studied polyelectrolytes, and the modifying monolayers provide the value of the polystyrene/water interfacial energy, which indicates the potential biocompatibility of the material based on the modi ed polystyrene.

Accurate computational design of three-dimensional protein crystals.

Protein crystallization plays a central role in structural biology. Despite this, the process of crystallization remains poorly understood and highly empirical, with crystal contacts, lattice packing arrangements and space group preferences being largely unpredictable. Programming protein crystallization through precisely engineered side-chain-side-chain interactions across protein-protein interfaces is an outstanding challenge. Here we develop a general computational approach for designing three-dimensional protein crystals with prespecified lattice architectures at atomic accuracy that hierarchically constrains the overall number of degrees of freedom of the system. We design three pairs of oligomers that can be individually purified, and upon mixing, spontaneously self-assemble into >100 µm three-dimensional crystals. The structures of these crystals are nearly identical to the computational design models, closely corresponding in both overall architecture and the specific protein-protein interactions. The dimensions of the crystal unit cell can be systematically redesigned while retaining the space group symmetry and overall architecture, and the crystals are extremely porous and highly stable. Our approach enables the computational design of protein crystals with high accuracy, and the designed protein crystals, which have both structural and assembly information encoded in their primary sequences, provide a powerful platform for biological materials engineering.

Dual regulation of crystal structure and purity of Ti2SC in the Ti/TiC/FeS2 system via enhancing reactant contact and controlling crystal plane growth rates

Typically, the X-ray diffraction spectrum of a highly oriented structure (HOS) MAX phase displays a prominent (002) characteristic peak. Surprisingly, this principle does not hold true for Ti2SC. Instead, the synthesized Ti2SC consistently exhibits a polyhedral structure (PS). Furthermore, even with hot-pressing sintering, the purity of Ti2SC remains relatively low. The primary factors contributing to anomalous crystal structures and low purity are insufficient crystal growth and limited reactant contact. In this study, we successfully achieved dual regulation of the crystal structure and purity of Ti2SC within the Ti/TiC/FeS2 system, which was accomplished by enhancing reactant contact and controlling the growth rates of crystal planes. Notably, the intermediate products FeS and Fe, existing in a liquid phase at varying temperatures, played a crucial role in both the formation of Ti2SC and the growth of Ti2SC crystals. As a result, PS-Ti2SC and HOS–Ti2SC with purity greater than 99 wt% were successfully prepared. The formation of PS-Ti2SC can be attributed to the inadequate growth of each crystal plane at relatively lower temperatures (1500 °C). While the synthesis of HOS–Ti2SC necessitates the utilization of both a liquid phase and high temperature, as the epitaxial growth mechanism followed by vertical growth has been demonstrated to facilitate the generation of highly oriented crystals. The liquid phase facilitates the swift growth of Ti2SC crystals along the (001) crystal plane, while the (002) crystal plane necessitates a higher temperature (1760 °C) for relatively rapid growth. This work will open a new pathway for the development of the MAX phase from the perspective of crystal structure regulation and high purification preparation.

Crystal violet adsorption onto modified biosorbent prepared from agricultural waste: kinetics, isotherm and thermodynamic studies

The present study investigated crystal violet removal by modifying the chestnut shell with a chemical activation method. For this purpose, H2SO4 and NaOH pretreatments were applied to the chestnut shell and the pretreatment method that gave the best performance under the same conditions was determined. The best adsorption efficiency was achieved with the NaOH pretreatment (99.06%) and the crystal violet adsorption reached equilibrium within 60 min. After selecting the best chemical activation method, the modified chestnut shell was characterized before and after adsorption (FTIR and SEM). Furthermore, the effects of parameters such as pH, initial crystal violet concentration, adsorbent dosage, temperature, and contact time were observed. Moreover, isotherm, kinetics and thermodynamics of the adsorption process were researched in detail. The best results were obtained with the Langmuir isotherm model (R2=0.99) and the pseudo-second-order kinetic model (R2=0.99). Thermodynamic parameters showed that the adsorption process is spontaneous, endothermic and feasible.

Fusion crystallization reveals the behavior of both the 1TEL crystallization chaperone and the TNK1 UBA domain

Human thirty-eight-negative kinase-1 (TNK1) is implicated in cancer progression. The TNK1 ubiquitin-associated (UBA) domain binds polyubiquitin and plays a regulatory role in TNK1 activity and stability. No experimentally determined molecular structure of this unusual UBA domain is available. We fused the UBA domain to the 1TEL variant of the translocation ETS leukemia protein sterile alpha motif (TELSAM) crystallization chaperone and obtained crystals diffracting as far as 1.53Å. GG and GSGG linkers allowed the UBA to reproducibly find a productive binding mode against its host 1TEL polymer and crystallize at protein concentrations as low as 0.2mg/mL. Our studies support a mechanism of 1TEL fusion crystallization and show that 1TEL fusion crystals require fewer crystal contacts than traditional protein crystals. Modeling and experimental validation suggest the UBA domain may be selective for both the length and linkages of polyubiquitin chains.