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Time-Resolved Serial Femtosecond Crystallography (TR-SFX) conducted at X-ray Free Electron Lasers (XFELs) has become a powerful tool for capturing macromolecular structural movies of light-initiated processes. As the capabilities of XFELs advance, we anticipate that a new range of coherent control and structural Raman measurements will become achievable. Shorter optical and x-ray pulse durations and increasingly more exotic pulse regimes are becoming available at free electron lasers. Moreover, with high repetition enabled by the superconducting technology of European XFEL (EuXFEL) and Linac Coherent Light Source (LCLS-II) , it will be possible to improve the signal-to-noise ratio of the light-induced differences, allowing for the observation of vibronic motion on the sub-Angstrom level. To predict and assign this coherent motion, which is measurable with a structural technique, new theoretical approaches must be developed. In this paper, we present a theoretical density matrix approach to model the various population and coherent dynamics of a system, which considers molecular system parameters and excitation conditions. We emphasize the use of the Wigner transform of the time-dependent density matrix, which provides a phase space representation that can be directly compared to the experimental positional displacements measured in a TR-SFX experiment. Here, we extend the results from simple models to include more realistic schemes that include large relaxation terms. We explore a variety of pulse schemes using multiple model systems using realistic parameters. An open-source software package is provided to perform the density matrix simulation and Wigner transformations. The open-source software allows us to define any arbitrary level schemes as well as any arbitrary electric field in the interaction Hamiltonian. [ABSTRACT FROM AUTHOR]

The structure and mechanism of the water-oxidation chemistry that occurs in photosystem II have been subjects of great interest. The advent of X-ray free electron lasers allowed the determination of structures of the stable intermediate states and of steps in the transitions between these intermediate states, bringing a new perspective to this field. The room-temperature structures collected as the photosynthetic water oxidation reaction proceeds in real time have provided important novel insights into the structural changes and the mechanism of the water oxidation reaction. The time-resolved measurements have also given us a view of how this reaction-which involves multielectron, multiproton processes-is facilitated by the interaction of the ligands and the protein residues in the oxygen-evolving complex. These structures have also provided a picture of the dynamics occurring in the channels within photosystem II that are involved in the transport of the substrate water to the catalytic center and protons to the bulk.

Monoclonal antibodies (mAbs) that target the P-amyloid peptide (Aβ) are important Alzheimer's disease research tools and are now being used as Alzheimer's disease therapies. Conformation-specific mAbs that target oligomeric and fibrillar Aβ assemblies are of particular interest, as these assemblies are associated with Alzheimer's disease pathogenesis and progression. This paper reports the generation of rabbit mAbs against two different triangular trimers derived from Aβ. These antibodies are the first mAbs generated against Aβ oligomer mimics in which the high-resolution structures of the oligomers are known. We describe the isolation of the mAbs using single B-cell sorting of peripheral blood mononuclear cells (PBMCs) from immunized rabbits, the selectivity of the mAbs for the triangular trimers, the immunoreactivity of the mAbs with aggregated Aβ42, and the immunoreactivity of the mAbs in brain tissue from the 5xFAD Alzheimer's disease mouse model. The characterization of these mAbs against structurally defined trimers derived from Aβ enhances understanding of antibody-amyloid recognition and may benefit the development of diagnostics and immunotherapies in Alzheimer's disease.

In their folded state, biomolecules exchange between multiple conformational states that are crucial for their function. Traditional structural biology methods, such as X-ray crystallography and cryogenic electron microscopy (cryo-EM), produce density maps that are ensemble averages, reflecting molecules in various conformations. Yet, most models derived from these maps explicitly represent only a single conformation, overlooking the complexity of biomolecular structures. To accurately reflect the diversity of biomolecular forms, there is a pressing need to shift toward modeling structural ensembles that mirror the experimental data. However, the challenge of distinguishing signal from noise complicates manual efforts to create these models. In response, we introduce the latest enhancements to qFit, an automated computational strategy designed to incorporate protein conformational heterogeneity into models built into density maps. These algorithmic improvements in qFit are substantiated by superior Rfree and geometry metrics across a wide range of proteins. Importantly, unlike more complex multicopy ensemble models, the multiconformer models produced by qFit can be manually modified in most major model building software (e.g., Coot) and fit can be further improved by refinement using standard pipelines (e.g., Phenix, Refmac, Buster). By reducing the barrier of creating multiconformer models, qFit can foster the development of new hypotheses about the relationship between macromolecular conformational dynamics and function.

We report the synthesis, characterization, anti-cancer activity and mechanism of action of a novel water-soluble Cu(II) complex with salicylidene carbohydrazide as the ligand and o-phenanthroline as the co-ligand. The synthesized complex (1) was characterized by FT-IR, EPR, and electronic spectroscopy, as well as single crystal X-ray diffraction. This compound was found to be paramagnetic from EPR spectra and X-ray crystallography revealed that the molecule crystallized in an orthorhombic crystal system. The crystal lattice was asymmetric containing two distinct binuclear copper complexes containing the Schiff base as the major ligand, o-phenanthroline as a co-ligand, two nitrate anions, and two water molecules. The Cu(II) in the first site coordinated with the enolised ligand comprising enolate O-, phenolate O-, and the imine N and N,N from o-phen. The major part of this complex exists as Cu(II) coordinated with two H2O molecules at the second site with nitrate acting as the counter anion. However, a smaller portion of the complex exists where Cu(II) is coordinated with NO3- and H2O, and the remaining water molecule acts as lattice water. It was tested for DNA binding and cleavage properties which revealed that it binds in an intercalative mode to CT-DNA with Kb value of 1.25 x 104 M-1. Furthermore, cleavage studies reveal that the complex has potential for efficient DNA cleavage under both oxidative and hydrolytic conditions. It was able to enhance the rate of cleavage by 2.8 x 108 times. The complex shows good cytotoxicity to breast cancer monolayer (2D) as well as spheroid (3D) systems. The IC50 values for MDA-MB-231 and MCF-7 monolayer culture was calculated as 1.86 ± 0.17 µM and 2.22 ± 0.08 µM, respectively, and in (3D) spheroids of MDA-MB-231 cells, the IC50 value was calculated to be 1.51 ± 0.29 µM. It was observed that the complex outperformed cisplatin in both breast cancer cell lines. The cells treated with complex 1 underwent severe DNA damage, increased oxidative stress and cell cycle arrest which finally led to programmed cell death or apoptosis in triple negative breast cancer cells through an intrinsic pathway. [ABSTRACT FROM AUTHOR]

In this study, the chemoselective synthesis of two new thiophene derivatives is presented. The structure of newly synthesized thiophenes derivatives; ethyl 4-acetyl-3-phenyl-5-(phenylamino)thiophene-2-carboxylate (5) and ethyl (E)-4-(3-(dimethylamino)acryloyl)-3-phenyl-5-(phenylamino)thiophene-2-carboxylate (8) were established using different FTIR and NMR spectral analyses. Compound 8 was isolated as single crystal and its 3D structure was determined using X-ray crystallographic analysis. Possible intermolecular interactions that control the molecular packing of 8 were elucidated using Hirshfeld topology analysis. The O...H (13.7%), H...H (55.3%) and C...C (2.3%) intermolecular interactions are the most significant. Fukui functions showed that C4 in thiophene 5 and C3 in thiophene 8 are the most reactive atoms for nucleophilic attack, while N9 in thiophene 5 and C1 in thiophene 8 are the most reactive atoms for electrophilic attack. Antitumor activity of thiophene 5 was assessed and the results showed higher activity against HepG-2 (7.46 µg/mL) compared to the HCT 116 (12.60 µg/mL) cell line. [ABSTRACT FROM AUTHOR]

Light chain amyloidosis (AL), is classified as a plasma cell dyscrasia, whereby a mutant plasma cell multiplies uncontrollably and secretes enormous amounts of immunoglobulin‐free light chain (FLC) fragments. These FLCs undergo a process of misfolding and aggregation into amyloid fibrils, that can cause irreversible system‐wide damage. Current treatments that focus on depleting the underlying plasma cell clone are often poorly tolerated, particularly in patients with severe cardiac involvement, meaning patient prognosis is poor. An alternative treatment approach currently being explored is the inhibition of FLC aggregation by stabilisation of the native conformer. Here, we aimed to identify and characterise antibody fragments that target FLC domains and promote their stabilisation. Using phage‐display screening methods, we identified a variable heavy (VH) domain, termed VH1, targeted towards the FLC. Using differential scanning fluorimetry and surface plasmon resonance, VH1 was characterised to bind and kinetically stabilise an amyloidogenic FLC, whereby a > 5.5 °C increase in thermal stability was noted. This improved stability corresponded to the inhibition of fibril formation, where 10 : 1 LC : VH1 concentration reduced aggregation to baseline levels. X‐ray crystallographic structures of the LC : VH1 complex at atomic resolution revealed binding in a 1 : 1 ratio, mimicking the dimeric antigen binding sites of the native immunoglobulin molecule and the native LC homodimer. [ABSTRACT FROM AUTHOR]

Crystallization of the antimicrobial compound furacin (nitrofurazone) from 96%, 50%, and 20% hydrogen peroxide (HP) led to three novel solvates C6H6N4O4·H2O2, C6H6N4O4·1.5(H2O2), and C6H6N4O4·3.5(H2O2), respectively. Surprisingly, solvatomorphs that were richer in hydrogen peroxide were obtained from more dilute H2O2 solutions. The stability of the compounds decreased with an increase in peroxide content. These compounds represent a rare example of solvatomorphism (peroxomorphism) among crystalline hydrogen peroxide adducts. The structures of all the adducts were determined using X-ray crystallography. All of them contain a 1 : 1 chelating supramolecular synthon with a strong and unusual bifurcated donor hydrogen bond HOOH⋯(O2N, furane). Compound C6H6N4O4·3.5(H2O2) is the first example of a structure containing 2D hydrogen-bonded layers built exclusively from peroxide molecules. In its crystal, a concerted flip-flop proton disorder of HP molecules was observed for the first time. [ABSTRACT FROM AUTHOR]

Six Schiff bases with general name 5-(diethylamino)-2-(((halophenyl)imino)methyl)phenol (where halo = 4-fluoro (H1), 2-fluoro (H2), 2-bromo (H3), 4-bromo (H4), 4-chloro (H5) and 3-chloro-4-fluoro (H6)) were prepared by the condensation reaction between 4-(diethylamino)salicylaldehyde and suitable halogenated aromatic amines. The six halogenated Schiff bases were elucidated using different spectroscopic techniques and the structure of H3 and H6 were confirmed using single-crystal X-ray crystallography. The bond lengths of C7–N1, C7–C8 and C8–C9 obtained from structural analysis for both compounds depicted their enol-tautomeric characteristic form. The Hirshfeld analysis revealed that H‧‧‧H intermolecular contacts contributed most towards the Hirshfeld surfaces of both H3 (47.6%) and H6 (39.9%). Quantum chemical calculation studies showed that H1 and H2 have the highest and lowest energy band gap (∆E = 3.80 eV for H1 and ∆E = 3.73 eV for H2), depicting H2 and H1 as the most and least chemically reactive respectively among all the compounds. α-Amylase and α-glucosidase assay were used to evaluate the antidiabetes prowess of the synthesized compounds. All the compounds exhibited lower IC50 values than acarbose (reference drug) in α-amylase assay experiments and H5 with lowest IC50 value of 63.54 μM could be suggested to have the highest α-amylase inhibitory potential among the test samples. For α-glucosidase assay, H1–H6 displayed good antidiabetic potential. However, none of the compounds outshined acarbose with H6 having highest α-glucosidase inhibitory potential when compared to others i.e., IC50 of H6 = 60.89 μM and IC50 of acarbose = 51.42 μM. We measured the antioxidant potential of H1–H6 exploring 2,2-diphenyl-1-picrylhydrazyl (DPPH), nitric oxide (NO) and ferric reducing ability power (FRAP) assays. The DPPH as well as NO radical scavenging assay showed that all the compounds exhibited excellent antioxidant results with some of the compounds surpassing catechin (reference drug). Compound H5 with IC50 values of 30.32 mM and 31.73 mM outshined catechin with IC50 values of 31.17 mM and 140.62 mM for DPPH and NO assays respectively. All the compounds fell within the threshold of Lipinski's Ro5 projecting them as orally bioavailable and less toxic future therapeutics. [ABSTRACT FROM AUTHOR]

Microtubule-targeting agents are an important class of anti-cancer drugs; their full potential is however not realized because of significant myelotoxicity and neurotoxicity. We here report 3-nitropyridine analogues as a novel group of microtubule-targeting agents with potent anti-cancer effects against a broad range of cancer types. We show that these 3-nitropyridines induce cell cycle arrest in the G2-M phase and inhibit tubulin polymerization by interacting with tubulin. Determination of the tubulin–4AZA2996 structure by X-ray crystallography demonstrated that this class of compounds binds to the colchicine-site of tubulin. Furthermore, the anti-cancer effect was demonstrated both in vitro and in vivo in a murine heterotopic xenograft model of colon cancer. When administered intravenously, 4AZA2891 effectively inhibited cancer growth. Whereas 3-nitropyridine compounds do not induce myelotoxicity at pharmacological doses, the neurotoxicity associated with microtubule-targeting agents is still present. [ABSTRACT FROM AUTHOR]

PINK1 and Parkin mutations lead to the early onset of Parkinson's disease. PINK1-mediated phosphorylation of ubiquitin (Ub), ubiquitin-like protein (NEDD8), and ubiquitin-like (Ubl) domain of Parkin activate autoinhibited Parkin E3 ligase. The mechanism of various phospho-Ubls' specificity and conformational changes leading to Parkin activation remain elusive. Herein, we show that compared to Ub, NEDD8 is a more robust binder and activator of Parkin. Structures and biophysical/biochemical data reveal specific recognition and underlying mechanisms of pUb/pNEDD8 and pUbl domain binding to the RING1 and RING0 domains, respectively. Also, pUb/pNEDD8 binding in the RING1 pocket promotes allosteric conformational changes in Parkin's catalytic domain (RING2), leading to Parkin activation. Furthermore, Parkinson's disease mutation K211N in the RING0 domain was believed to perturb Parkin activation due to loss of pUb binding. However, our data reveal allosteric conformational changes due to N211 that lock RING2 with RING0 to inhibit Parkin activity without disrupting pNEDD8/pUb binding. [Display omitted] • pNEDD8/NEDD8 shows a higher affinity and activation of Parkin compared to pUb/Ub • Mechanism of specific binding of pUb/pNEDD8/pUbl to Parkin is revealed • Mechanism of pUb/pNEDD8 mediated Parkin activation is revealed • Mechanism of pathogenic mutation K211N mediated loss of Parkin activity is identified Lenka et al. determined the Parkin-pNEDD8 complex structure that reveals the specific recognition of Parkin activators on its two similar basic patches despite the similarities of various ubiquitin-like domains/proteins. They also explain the mechanism behind the reduced Parkin activity of the disease mutant K211N, which is of interest for drug discovery. [ABSTRACT FROM AUTHOR]

Non-heme iron-dependent sulfoxide/selenoxide synthases (NHISS) constitute a unique metalloenzyme class capable of installing a C–S/Se bond onto histidine to generate thio/selenoimidazole antioxidants, such as ergothioneine and ovothiol. These natural products are increasingly recognized for their health benefits. Among associated ergothioneine-biosynthetic enzymes, type IV EgtBs stand out, as they exhibit low sequence similarity with other EgtB subfamilies due to their recent divergence from the ovothiol-biosynthetic enzyme OvoA. Herein, we present crystal structures of two representative EgtB-IV enzymes, offering insights into the basis for this evolutionary convergence and enhancing our understanding of NHISS active site organization more broadly. The ability to interpret how key residues modulate substrate specificity and regioselectivity has implications for downstream identification of divergent reactivity within the NHISS family. To this end, we identify a previously unclassified clade of OvoA-like enzymes with a seemingly hybrid set of characteristics, suggesting they may represent an evolutionary intermediate between OvoA and EgtB-IV. [Display omitted] • Hercynine-bound structures offer insights into the convergent evolution of EgtB-IV • EgtB-IV cannot be converted into OvoA through simple active site mutations • An active site lid-loop dictates cysteine specificity in OvoA-like enzymes • A potential evolutionary intermediate between OvoA and EgtB-IV is presented Ireland et al. report X-ray crystal structures of type IV EgtB enzymes involved in ergothioneine biosynthesis. The results shed light on the convergent evolution of sulfoxide synthase EgtB-IV, which features closest homology to ovothiol-biosynthetic OvoA. Additionally, they clarify the structural basis for EgtB-IV's specificity for cysteine as the S-donor. [ABSTRACT FROM AUTHOR]

Schiff bases possess a range of unique physical, chemical and medicinal properties, which contribute to their potency as biological agents. A Schiff base was synthesized, which consisted of 4-chloro aniline and vanillin. The successful formation of the compound was confirmed by their comparable FTIR, UV–Vis, 1HNMR and 13CNMR spectra. The compound was analyzed using X-ray crystallography, which revealed that it crystallizes in the monoclinic system with the space group P21/c. The Hirshfeld surface analysis revealed valuable information about the intermolecular interactions present in the crystal lattice. The most common contacts observed were between hydrogen and hydrogen, as well as carbon and hydrogen. The obtained fluorescence values of 375 nm and 412 nm at excitation wavelength 322 nm strongly indicated the luminescent nature of the compound. The thermal stability of the compound was assessed through thermogravimetric analysis (TGA). DFT was used to optimize the geometry using the B3LYP/cc-pVDZ basis set. The compound exhibited an energy gap of 2.35 electron volts (eV). According to the analysis of molecular electrostatic potential, C and O atoms were found to be electrophilic. According to Mulliken charge analysis, C atoms possess both positive and negative charges, hydrogen atoms exclusively carry positive charges, and Cl, O and N atoms exclusively carry negative charges. Upon analysis of the molecule, the electron localization function discovered that the atoms of carbon, nitrogen and chlorine exhibited delocalized electron density. Also, the localized orbital locator identified the localized orbital positions in the hydrogen atoms. The reduced density gradient (RDG) maps exhibit a wide range of noncovalent interactions. In comparison to the standard amoxicillin, the synthesized compound demonstrated moderate antimicrobial efficacy against the gram-positive bacteria Klebsiella pneumoniae and the fungi Candida albicans. The compound’s concentration-dependent antioxidant and anti-inflammatory properties were demonstrated through in vitro biological evaluation using 2,2-diphenyl-1-picrylhydrazyl and human red blood cell (HRBC) methods. The compound was docked using the proteins 7BYE and 4LEB were chosen from both organisms. The lowest binding attractions obtained were -3.40 kcal/mol for 7BYE and -4.51 kcal/mol for 4LEB. To investigate the stability of the protein–ligand complex, molecular dynamic simulation was employed. The ligands in silico toxicity potentialities were analyzed using seven toxicity models and the results indicated that the ligand is biocompatible. [ABSTRACT FROM AUTHOR]

A novel rosarin di-Cu complex 2Cu-1 and a linear six-pyrrolic mono-copper complex 1Cu-1 were synthesized using rosarin as the ligand. The molecular conformations of these complexes were confirmed by X-ray crystallography. The optical study of 1Cu-1 indicated NIR-II absorption due to the long six-pyrrolic ligand and the ICT effect. The 2Cu-1 complex exhibited a very narrow electronic reduction–oxidation gap of 0.50 eV, attributed to the antiaromatic characteristics of the rosarin ring. The first HER study of the di-copper rosarin complex 2Cu-1 indicated that the multi-metal poly-pyrrolic complexes are promising molecular hydrogen evolution reaction catalysts. [ABSTRACT FROM AUTHOR]

Somatic mutations in the epidermal growth factor receptor (EGFR) are a major cause of non-small cell lung cancer. Among these structurally diverse alterations, exon 20 insertions represent a unique subset that rarely respond to EGFR tyrosine kinase inhibitors (TKIs). Therefore, there is a significant need to develop inhibitors that are active against this class of activating mutations. Here, we conducted biochemical analysis of the two most frequent exon 20 insertion variants, V769_D770insASV (insASV) and D770_N771insSVD (insSVD) to better understand their drug sensitivity and resistance. From kinetic studies, we found that EGFR insASV and insSVD are similarly active, but have lower Km, ATP values compared to the L858R variant, which contributes to their lack of sensitivity to 1st-3rd generation EGFR TKIs. Biochemical, structural, and cellular studies of a diverse panel of EGFR inhibitors revealed that the more recently developed compounds BAY-568, TAS6417, and TAK-788 inhibit EGFR insASV and insSVD in a mutant-selective manner, with BAY-568 being the most potent and selective versus wild-type (WT) EGFR. Cocrystal structures with WT EGFR reveal the binding modes of each of these inhibitors and of poziotinib, a potent but not mutantselective inhibitor, and together they define interactions shared by the mutant-selective agents. Collectively, our results show that these exon20 insertion variants are not inherently inhibitor resistant, rather they differ in their drug sensitivity from WT EGFR. However, they are similar to each other, indicating that a single inhibitor should be effective for several of the diverse exon 20 insertion variants. [ABSTRACT FROM AUTHOR]

In this study, we present a detailed analysis of the structural forms of asymmetric curcuminoids through single‐crystal X‐ray diffraction and nuclear magnetic resonance (NMR) spectroscopy. Curcuminoids substituted with electron‐withdrawing groups (Cl and Br) predominantly adopt the β‐keto‐enol form, whereas those with an OMe group exhibit both diketo and β‐keto‐enol forms. Variable‐temperature X‐ray diffraction studies confirm that the structural state of curcuminoids is influenced by temperature. Our findings also reveal that the β‐keto‐enol form is favored in acidic and neutral conditions, while the diketo form is more prevalent in basic environments. The theoretical Density Functional Theory (DFT) calculations further elucidate the energy differences between the diketo and β‐keto‐enol forms, emphasizing the significance of electronic effects and temperature on tautomeric distributions. These insights enhance our understanding of the structural dynamics of curcuminoids, with implications for their biological activity and pharmacological applications. [ABSTRACT FROM AUTHOR]

Allylic oxygenated derivatives of himachalenes are highly valued molecules due to their potential applications in perfumery, cosmetics, and pharmaceuticals. Previous attempts at catalyzed allylic oxidation of himachalenes led to the formation of a very stable η3-allyl palladium complex, preventing any further reaction development. Herein, we present the first successful palladium-catalyzed synthesis of a novel allylic acetoxylated derivative of himachalenes. This reaction was achieved by incorporating an aromatic ring into the substrate structure. The resulting intermediate complex was isolated and characterized using nuclear magnetic resonance spectroscopy and X-ray crystallography. Density functional theory (DFT) calculations were performed to compare the reactivity of the newly synthesized complex with previously reported ones. The theoretical results confirm that the introduction of an aromatic ring enhances the reactivity of the η³-allyl palladium complex, thereby facilitating the desired transformation. [ABSTRACT FROM AUTHOR]

The activation of inert C–H bonds remains a challenge in current chemistry. Here, we report the excellent reactivity of the anionic gallylene species [LGa:][Na(THF)3] (L = [(2,6-iPr2C6H3)NC(CH3)]22−, 1) that allows the selective activation one ortho sp2 C–H bond of several azobenzene and azide derivatives at ambient temperature, with the transfer of the hydrogen atom to one of the nitrogen atoms. The process leads to the formation of the aryl amido products [LGa-κ2N,C-PhNN(H)(p-R-C6H3)][Na(solvent)3] (2, R = H solvent = DME (1,2-Dimethoxyethane); 3, R = –OMe, solvent = DME; 4, R = –NMe2 solvent = THF), [LGa-κ2N,C-(m-CH3-C6H4)NN(H)(m-CH3-C6H3)][Na(15-C-5)2] (5) with new Ga–C and Ga–N bonds. Moreover, 1 is also effective for the C–H activation of two azides RN3 (R = 2,4,6-Me3C6H2 or 2,6-iPr2C6H3), resulting in the formation of gallium amides [LGa(NH-2-(CH2)-4,6-Me2C6H2)][Na(15-C-5)2] (6) and [LGa(NH-2,6-iPr2C6H3)2][Na(THF)5] (7) through intra- or intermolecular sp3 C–H amination. Significantly, these reactions occur for the highly challenging activation of inert C(sp2)–H and C(sp3)–H bonds, thus demonstrating the excellent reactivity of the Ga(I) species 1. The products 2–7 were characterized by X-ray crystallography, 1H and 13C NMR, UV–vis spectroscopy, and density functional theory (DFT) calculations. [ABSTRACT FROM AUTHOR]

• A metastable degradant of Cimlanod was isolated via LC-HRMS/MS guided synthesis. • Structure elucidation of the degradant was conducted via single crystal X-ray crystallography. • Plausible degradation pathways of Cimlanod in the prototype formulation solution under stressed conditions were proposed. N -hydroxy-5-methylfuran-2-sulfonamide (BMS-986231, Cimlanod) was being developed as a pH-sensitive prodrug of HNO (nitroxyl) for the treatment of acute decompensated heart failure. During a stressed study of Cimlanod in a prototype formulation solution (pH 4.5) at 40°C, a predominant unknown degradant along with three previously identified degradants were observed. The unknown degradant was isolated from the stressed solution via preparative HPLC but totally decomposed during freeze-drying. LC-HRMS analysis of the isolated unknown degradant, prior to freeze-drying, revealed an empirical formula equivalent to the adduct of Cimlanod with SO 2 even though SO 2 was not added in the prototype formulation solution. The unknown degradant was synthesized from Cimlanod and DABSO ((1,4-diazabiscyclo[2,2,2]octane bis(sulfur dioxide) adduct) and isolated as a crystalline DABCO (1,4-diazabiscyclo[2,2,2]octane) salt for single crystal X-ray structure elucidation. The degradation of Cimlanod increased when the solution was exposed to air, as compared to N 2 atmosphere. A plausible mechanism was postulated for the unexpected degradation pathway of Cimlanod. This study provided in-depth stability knowledge of Cimlanod, which will be beneficial to the subsequent stability indicating method development and validation as well as the registrational applications on the content and qualification of impurities in new drug products. [ABSTRACT FROM AUTHOR]

Serine protease inhibitor B9 (serpin B9, also known as protease inhibitor 9 or PI9) plays a critical role in regulating the immune response by specifically inhibiting granzyme B, a serine protease found in cytotoxic T lymphocytes and natural killer cells. Despite its potential as an anticancer drug target, the structural details of serpin B9 have remained elusive until now. In this study, a cleaved form of recombinant human serpin B9 was successfully prepared and crystallized. The crystals belonged to space group P212121, with unit‐cell parameters a = 68.51, b = 82.32, c = 101.17 Å, and an X‐ray diffraction data set was collected at 1.9 Å resolution. The structure shows that serpin B9 adopts a relaxed conformation, with its cleaved reactive‐centre loop inserted into the central β‐sheet. Unlike other serpins, serpin B9 shows significant structural deviations around helix D, with a larger surface cavity, which could serve as a promising target for small‐molecule inhibitors. [ABSTRACT FROM AUTHOR]

Anti‐immunocomplex (Anti‐IC) antibodies have been used in developing noncompetitive immunoassays for detecting small molecule analytics (haptens). These antibodies bind specifically to the primary antibody in complex with hapten. Although several anti‐IC antibody–based immunoassays have been developed, structural studies of these systems are very limited. In this study, we determined the crystal structures of anti‐testosterone Fab220 in complex with testosterone and the corresponding anti‐IC antibody FabB12. The structure of the ternary complex of testosterone, Fab220, and FabB12 was predicted using LightDock and AlphaFold. The ternary complex has a large (~ 1100 Å2) interface between antibodies. The A‐ring of the testosterone bound by Fab220 also participates in the binding of the anti‐IC antibody. The structural analysis was complemented by native mass spectrometry. The affinities for testosterone (TES) and three cross‐reactive steroids [dihydrotestosterone (DHT), androstenedione (A4), and dehydroepiandrosterone sulfate (DHEA‐S)] were measured, and ternary complex formation was studied. The results clearly show the ternary complex formation in the solution. Although DHT showed significant cross‐reactivity, A4 and DHEA‐S exhibited minor cross‐reactivity. [ABSTRACT FROM AUTHOR]

The biological denitrification process is crucial for removing nitrogen and reducing nitrous oxide (N2O) emissions. To investigate the mechanism of action of key enzymes in the denitrification process, it is necessary to analyze the protein structure, active site, and conformational changes of denitrifying enzymes. X-ray crystallography, nuclear magnetic resonance (NMR), and cryogenic electron microscopy (Cryo-EM), as well as spectroscopic analyses such as Raman resonance and UV-vis, have been used to study the structure of denitrifying enzymes. However, practical application is limited by protein folding and assembly, as well as issues of purity, concentration, and homogeneity. This review summarizes the biochemical properties of the four denitrifying enzymes and typical purification methods. Additionally, this review emphasizes important factors to consider during protein expression, crude extract, and chromatographic purification. It also provides insight into the future development of protein purification methods for denitrifying enzymes. This review aims to bridge the gap between the complex biochemical process of denitrifying enzymes and the technical intricacies of protein purification, with the ultimate goal of reducing N2O emissions in ecosystems. [ABSTRACT FROM AUTHOR]

Lignocellulolytic clostridia employ multiple pairs of alternative σ/anti‐σ (SigI/RsgI) factors to regulate cellulosomal components for substrate‐specific degradation of cellulosic biomass. The current model has proposed that RsgIs use a sensor domain to bind specific extracellular lignocellulosic components and activate cognate SigIs to initiate expression of corresponding cellulosomal enzyme genes, while expression of scaffoldins can be initiated by several different SigIs. Pseudobacteroides cellulosolvens contains the most complex known cellulosome system and the highest number of SigI–RsgI regulons yet discovered. However, the function of many RsgI sensor domains and their relationship with the various enzyme types are not fully understood. Here, we report that RsgI4 from P. cellulosolvens employs a C‐terminal module that bears distant similarity to the fibronectin type III (Fn3) domain and serves as the sensor domain. Substrate‐binding analysis revealed that the Fn3‐like domain of RsgI4 represents a novel carbohydrate‐binding module (CBM) that binds to a wide range of polysaccharide types. Structure determination further revealed that the Fn3‐like domain belongs to the type B group of CBMs with a predicted concave face for substrate binding. Promoter sequence analysis of cellulosomal genes revealed that SigI4 is responsible for cellulosomal regulation of major scaffoldins rather than enzymes, consistent with the broad substrate specificity of the RsgI4 sensor domain. Notably, scaffoldins are invariably required as cellulosome components regardless of the substrate type. These findings suggest that the intricate cellulosome system of P. cellulosolvens comprises a more elaborate regulation mechanism than other bacteria and thus expands the paradigm of cellulosome regulation. [ABSTRACT FROM AUTHOR]

The acetylpyrrole scaffold is an acetylated lysine mimic that has been previously explored to develop bromodomain inhibitors. When tested on the hepatoma cell line Huh7 and the breast cancer cell line MDA‐MB‐231, a few compounds in our acetylpyrrole‐thiazole library induced peculiar morphological changes, progressively causing cell death at increasing concentrations. Their evaluation on a panel of human bromodomains revealed concurrent inhibition of BRPF1 and BET bromodomains. To dissect the observed cellular effects, the acetylpyrrole derivatives were compared to JQ1 and GSK6853, chemical probes for the bromodomains of BET and BRPF1, respectively. The appearance of neurite‐like extrusions, accompanied by βIII‐tubulin overexpression, is caused by BET inhibition, with limited effect on cellular viability. Conversely, interference with BRPF1 induces cellular death but not phenotypic alterations. Combined treatment with JQ1 and GSK6853 showed additivity in reducing cellular viability, comparably to the acetylpyrrole‐thiazole‐based BET/BRPF1 inhibitors. In addition, we determined the crystallographic structures of the BRD4 and BRPF1 bromodomains in complex with the acetylpyrrole‐thiazole compounds. The binding modes in the two bromodomains show similar interactions for the acetylpyrrole and different orientations of the moiety that point to the rim of the acetyl‐lysine pocket. [ABSTRACT FROM AUTHOR]

Deoxyuridine triphosphate pyrophosphatase (DUT) suppresses incorporation of uracil into genomic DNA during replication. Thermostable DUTs from hyperthermophilic archaea such as Thermococcus pacificus enhance PCR amplification by preventing misincorporation of dUTP generated by spontaneous deamination of dCTP. However, it is necessary to elucidate whether DUTs do not cause dNTP imbalances during PCR by unwanted side activity. Moreover, it has been unknown what structural features define the thermostability of those DUTs. Here, DUT from a hyperthermophilic eubacterium, Aquifex aeolicus (Aa‐DUT), was characterized together with those from T. pacificus (Tp‐DUT). Aa‐DUT was as thermostable as Tp‐DUT up to at least 95°C. The crystal structures of the two thermostable enzymes were determined, which revealed that the structures of Aa‐DUT and Tp‐DUT resembled those of monofunctional and bifunctional DUTs, respectively. Generally, bifunctional DUTs harbor the dCTP deaminase activity in addition to the DUT activity. However, not only Aa‐DUT but also Tp‐DUT showed poor activity towards dCTP, indicating both enzymes are monofunctional. We further examined eight types of parameters related to thermostability of protein structure and found that the thermostability of Aa‐DUT and Tp‐DUT might be accomplished by increased numbers of ion pairs on the protein surface. Finally, we verified that Aa‐DUT promoted PCR amplification with Pfu DNA polymerase to the same extent as Tp‐DUT. Collectively, we conclude that both DUTs from hyperthermophiles maintain the enzymatic activity at high temperatures without consuming dCTP due to the lack of the deaminate activity. [ABSTRACT FROM AUTHOR]

Identifying epitopes, or the segments of a protein that bind to antibodies, is critical for the development of a variety of immunotherapeutics and diagnostics. In vaccine design, the intent is to identify the minimal epitope of an antigen that can elicit an immune response and avoid off-target effects. For prognostics and diagnostics, the epitope-antibody interaction is exploited to measure antigens associated with disease outcomes. Experimental methods such as X-ray crystallography, cryo-electron microscopy, and peptide arrays are used widely to map epitopes but vary in accuracy, throughput, cost, and feasibility. By comparing machine learning epitope mapping tools, we discuss the importance of data selection, feature design, and algorithm choice in determining the specificity and prediction accuracy of an algorithm. This review discusses limitations of current methods and the potential for machine learning to deepen interpretation and increase feasibility of these methods. We also propose how machine learning can be employed to refine epitope prediction to address the apparent promiscuity of polyreactive antibodies and the challenge of defining conformational epitopes. We highlight the impact of machine learning on our current understanding of epitopes and its potential to guide the design of therapeutic interventions with more predictable outcomes. [ABSTRACT FROM AUTHOR]

ABSTRACT In this study, we report the synthesis, characterization, and biological investigation of antimony(III) halide complexes with various coordination architectures constructed from thiophene thiosemicarbazones. Antimony(III) thiophene‐2‐carbaldehyde thiosemicarbazone complexes (1, 2, 4, and 5) exhibit a square pyramidal geometry, with ligands coordinated to the central antimony atom in two distinct binding modes. In contrast, antimony(III) 2‐acetylthiophene thiosemicarbazone complexes (3, 6, and 7) adopt a seesaw geometry. These complexes (1–7) represent the first reported examples of antimony(III) halide thiosemicarbazone compounds. The unique coordination environments observed in these complexes are of significant importance within the realm of antimony chemistry. These synthesized complexes exhibit different coordination geometries as well as potential biological activities. The antiproliferative activity against the human breast adenocarcinoma (MCF‐7) cell line and antimicrobial activity against Gram‐positive and Gram‐negative bacteria were assessed. Antimony(III) thiophene‐2‐carbaldehyde thiosemicarbazone complexes (1, 2, 4, and 5) showed significant antiproliferative activity with IC50 values ranging from 8.5 to 19.1 μM, while antimony(III) 2‐acetylthiophene thiosemicarbazone complexes (3, 6, and 7) had higher IC50 values. Additionally, the antimony complexes demonstrated selective antimicrobial activity against Gram‐negative bacteria. [ABSTRACT FROM AUTHOR]

Metals have been used in medicine for centuries. However, it was not until much later that the effects of inorganic drugs could be rationalized from a mechanistic point of view. Today, thanks to the technologies available, this approach has been functionally developed and implemented. It has been found that there is probably no single biological target for the pharmacological effects of most inorganic drugs. Herein, we present an overview of some integrated and multi‐technique approaches to elucidate the molecular interactions underlying the biological effects of metallodrugs. On this premise, selected examples are used to illustrate how the information obtained on metal‐based drugs and their respective mechanisms can become relevant for applications in fields other than medicine. For example, some well‐known metallodrugs, which have been shown to bind specific amino acid residues of proteins, can be used to solve problems related to protein structure elucidation in crystallographic studies. Diruthenium tetraacetate can be used to catalyze the conversion of hydroxylamines to nitrones with a high selectivity when bound to lysozyme. Finally, a case study is presented in which an unprecedented palladium/arsenic‐mediated catalytic cycle for nitrile hydration was discovered thanks to previous studies on the solution chemistry of the anticancer compound arsenoplatin‐1 (AP‐1). [ABSTRACT FROM AUTHOR]

Psilocybin, the natural hallucinogen from Psilocybe (magic) mushrooms, is a highly promising drug candidate for the treatment of depression and several other mental health conditions. Biosynthesis of psilocybin from the amino acid l‐tryptophan involves four strictly sequential modifications. The third of these, ATP‐dependent phosphorylation of the intermediate 4‐hydroxytryptamine, is catalysed by PsiK. Here we present a crystallographic analysis and a structure‐based mutagenesis study of this kinase, providing insight into its mode of substrate recognition. The results of our work will support future bioengineering efforts aimed at generating variants of psilocybin with enhanced therapeutic properties. [ABSTRACT FROM AUTHOR]

Pyrrolizidine alkaloids (PAs) are a structurally diverse group of heterocyclic specialized metabolites characterized by a core structure comprising a hexahydro‐1H‐pyrrolizine. PAs are synthesized through two main pathways. In plants, assembly occurs via a homospermidine synthase, and in bacteria, through combined action of a nonribosomal peptide synthetase and a Baeyer–Villiger monooxygenase. While the toxic properties of plant‐derived PAs and their prevalence in animal and human foods have been extensively studied, the biological roles and biosynthesis of more complex bacterial PAs are not well understood. Here, we report the identification and characterization of a bacterial biosynthetic gene cluster from Xenorhabdus hominickii, xhpA‐G, which is responsible for producing the PA pseudo‐dimer pyrrolizwilline. Analysis of X. hominickii promoter exchange mutants together with heterologous expression of xhpA‐G in E. coli, revealed a set of pathway intermediates, two of which were chemically synthesized, as well as multiple derivatives. This information was leveraged to propose a detailed biosynthetic pathway to pyrrolizwilline. Furthermore, we have characterized the hydrolase XhpG, the key enzyme in the conversion of the pathway intermediate pyrrolizixenamide to pyrrolizwilline, using X‐ray crystallography and small‐angle X‐ray scattering (SAXS). [ABSTRACT FROM AUTHOR]

The strengths of H-bonding interactions in networks are affected by cooperativity between the interacting sites. Compounds with an intramolecular H-bond between a sulfonamide NH group and pyridine nitrogen were used to measure the magnitude of cooperative effects on intermolecular H-bonding interactions with the sulfonamide oxygen. X-ray crystallography and 1H NMR experiments confirm the presence of the intramolecular H-bond and show that it is maintained in the 1 : 1 complex formed with perfluoro-tert-butanol (PFTB) in n-octane solution. Association constants for formation of 1 : 1 complexes with PFTB were determined using UV/Vis absorption titrations for a series of compounds equipped with different pyridine groups. Substituents on the pyridine were used to tune the strength of the intramolecular H-bond and investigate the effects on the strength of the intermolecular H-bond. Electron-donating groups on the pyridine that increase the strength of the intramolecular H-bond were found to increase in the strength of the intermolecular interaction with PFTB. The results were used to determine the H-bond acceptor parameters, β, for the sulfonamide oxygen group, and the values show a linear relationship with the value of β for the pyridine nitrogen. The slope of this relationship corresponds to the cooperativity parameter, κ, which is +0.16. The positive cooperativity observed in H-bonded sulfonamides is comparable to the value measured previously for the amide group (κ = +0.20). [ABSTRACT FROM AUTHOR]

Terrestrial organisms have systems to escape from desiccation stresses. For example, tardigrades (also known as water bears) can survive severe dried and other extreme environments by anhydrobiosis. Although their extraordinary ability has enchanted people, little is known about the detailed molecular mechanisms of anhydrobiosis. Here, we focused on the tardigrade Ramazzottius varieornatus, one of the toughest animals on Earth. A transcriptome database of R. varieornatus shows that genes encoding a Ferritin‐like protein are upregulated during desiccation or ultraviolet radiation. This protein shows sequence similarity to enigmatic proteins in desiccation‐tolerant bacteria and plants, which are hypothesized to be desiccation‐related. However, because these proteins lack detailed biological information, their functions are relatively unknown. We determined an atomic (1.05 Å) resolution crystal structure of a Ferritin‐like protein from R. varieornatus. The structure revealed a dinuclear metal binding site, and we showed that this Ferritin‐like protein has phosphatase activity toward several metabolite compounds including unusual nucleotide phosphates produced by oxidative or radiation damage. We also found that a homologous protein from a desiccation‐ and ultraviolet‐tolerant bacterium Deinococcus radiodurans is a metabolite phosphatase. Our results indicate that through cleaning up damaged metabolites or regulation of metabolite levels, this phosphatase family can contribute to stress tolerances. This study provides a clue to one of the universal molecular bases of desiccation‐stress tolerance. [ABSTRACT FROM AUTHOR]

The exploration of artificial metal-peptide assemblies (MPAs) is one of the most exciting fields because of their great potential for simulating the dynamics and functionality of natural proteins. However, unfavorable enthalpy changes make forming discrete complexes with large and adaptable cavities from flexible peptide ligands challenging. Here, we present a strategy integrating metal-cluster building blocks and peptides to create chiral metal-peptide assemblies and get a family of enantiopure [R-/S-Ni3L2]n (n = 2, 3, 6) MPAs, including the R-/S-Ni6L4 capsule, the S-Ni9L6 trigonal prism, and the R-/S-Ni18L12 octahedron cage. X-ray crystallography shows MPA formation reactions are highly solvent-condition-dependent, resulting in significant changes in ligand conformation and discrete cavity sizes. Moreover, we demonstrate that a structure transformation from Ni18L12 to Ni9L6 in the presence of benzopyrone molecules depends on the peptide conformational selection in crystallization. This work reveals that a metal-cluster building block approach enables facile bottom-up construction of artificial metal-peptide assemblies. The use of metal clusters to construct artificial protein-mimic structures with adaptable cavities has potential for simulating the dynamics and functionality of natural proteins. Here, the authors develop a family of chiral metal-peptide assemblies using {Ni3} clusters and flexible peptides, resulting in structures such as octahedral cages, trigonal prisms, and capsules. [ABSTRACT FROM AUTHOR]

The synthesis and characterization of the lithium m‐terphenylhydridoborates [Li(2,6‐Mes2C6H3)BH3]2 (1), Li(2,6‐Mes2C6H3)2BH2 (2) and Li(OEt2)(2,6‐Mes2C6H3)2BH2 (3) are reported. Hirshfeld Atom Refinement (HAR) of the experimentally obtained molecular structures by single‐crystal X‐ray crystallography allowed the determination of the exact positions of the hydrogen atoms. The bond situations of the various B−H⋅⋅⋅Li linkages were investigated by a complementary bonding analysis using various methods including atoms in molecules (AIM), electron localizability indicator (ELI−D), non‐covalent interaction (NCI) index and the compliance matrix. [ABSTRACT FROM AUTHOR]

Calix[n]porphyrinoids containing both sp2 and sp3meso‐carbons can bind metals like porphyrinoids and anions like calix[n]pyrroles. Thus, calix[n]porphyrinoids are hybrid macrocycles of porphyrinoids and calix[n]pyrroles. The core‐modified calix[n]porphyrinoids resulted by replacing one or more pyrroles of calix[n]porphyrinoids with different heterocycles or carbacycles that possesses very attractive structural, spectral, and coordination properties. For the past two decades, several different types of core‐modified calix[n]porphyrinoids in which one or more pyrroles were replaced with other heterocycles such as thiophene, selenophene, tellurophene, silole, phosphole, pyridine, etc and carbacycles such as m/p‐phenylene have been synthesized and explored their structure, metal coordination and anion sensing properties. This review describes different synthetic strategies used for the synthesis of various types of core‐modified calix[4]phyrins and expanded calix[n]phyrins and discusses their key structural aspects, properties, coordination chemistry and applications. [ABSTRACT FROM AUTHOR]

ABSTRACT A thiophosphoric triamide (SPT) ligand P(S)[NH‐2Py]3 was synthesized and utilized as a ligand to make novel binuclear and mononuclear Co2+ and Cd2+ complexes. The compounds are the first examples of discrete chelate SPT complexes with a [N]3P(S)‐based backbone in which the SPT acts as a flexible tridentate N,N,N‐donor or N,N,O‐donor ligand. The metal cation features a hexacoordinate environment M(N)3(Cl)3/M(N)4(O)2 adopting a distorted octahedral geometry. Hirshfeld surface analysis affirms that the pyridinyl nitrogen is a more dominant H‐bond acceptor than the thiophosphoryl sulfur atom. The inhibitory activity against the MDA‐MB‐231 cancer cell line in vitro indicated a good inhibitory effect for all compounds. The mononuclear Co2+ complex showed a stronger inhibitory activity against cancer cells than other studied compounds with an IC50 of 62 ± 1.4 nM. Antioxidant and antihemolytic activity of the compounds showed appreciable performance with the highest activities for the Cd2+ complex. A computational DNA binding study by molecular docking indicated a good affinity of the compounds with the DNA receptor by a mixed mode of interactions. Docking was also performed on SARS‐CoV‐2 and monkeypox (Mpox), resulting in binding energies (−5.5 to −8.9 kcal/mol) that are competitive with current effective drugs. [ABSTRACT FROM AUTHOR]

Alkali–metal nickelates are a class of highly reactive heterobimetallic complexes derived from Ni(0)–olefins and polar organo-alkali–metal reagents. First reported over 50 years ago, it is only in recent years that these overlooked complexes have found formidable roles in sustainable catalysis and beyond. In this article, we will showcase the emerging catalytic applications of lithium nickelates and discuss the mechanisms by which these heterobimetallic complexes facilitate challenging cross-coupling reactions. We will also review the unique structure and bonding of alkali–metal nickelates, as interrogated by X-ray crystallography and complementary bonding analysis, and finally explore the diverse coordination and co-complexation chemistry of these heterobimetallic complexes. [ABSTRACT FROM AUTHOR]

Metal ions have important roles in supporting the catalytic activity of DNA-regulating enzymes such as topoisomerases (topos). Bacterial type II topos, gyrases and topo IV, are primary drug targets for fluoroquinolones, a class of clinically relevant antibacterials requiring metal ions for efficient drug binding. While the presence of metal ions in topos has been elucidated in biochemical studies, accurate location and assignment of metal ions in structural studies have historically posed significant challenges. Recent advances in X-ray crystallography address these limitations by extending the experimental capabilities into the long-wavelength range, exploiting the anomalous contrast from light elements of biological relevance. This breakthrough enables us to confirm experimentally the locations of Mg2+ in the fluoroquinolone-stabilized Streptococcus pneumoniae topo IV complex. Moreover, we can unambiguously identify the presence of K+ and Cl-ions in the complex with one pair of K+ ions functioning as an additional intersubunit bridge. Overall, our data extend current knowledge on the functional and structural roles of metal ions in type II topos. [ABSTRACT FROM AUTHOR]

AbstractNaphthalene and its derivatives are often determined as markers of environmental pollution, although they are also recognized as crucial building blocks in many beneficial compounds. In the course of this study, a range of 1,2-disubstituted naphthalenes bearing nitro, amine, sulfonate, chlorine, and hydroxyl substituents were synthesized and characterized using several spectroscopic techniques, including FT-IR,1H NMR,13C NMR, DEPT-90, DEPT-135, DEPTq, elemental analysis, and high-resolution mass (HRMs). The NMR analysis of the aromatic hydrocarbon moiety in these compounds emphasized the impact of substituent variations on the appearance of 1H and 13C NMR signals. To resolve the issue of overlapping one-dimensional (1D) 1H NMR signals and determine the connectivity of hydrogen and carbon atoms from the aromatic naphtayl ring, we employed a “two-dimensional” experiment, using homonuclear correlation spectroscopy (2D 1H-1H COSY) and heteronuclear single quantum correlation (2D 1H - 13C HSQC) and heteronuclear multiple‐bond correlation spectroscopy (2D 1H - 13C HMBC). This work also investigated the correlation between 1H NMR signals and thin-layer chromatography (TLC) data. X-ray crystallography data validates that the presence of a specific group, not aligned coplanarly with the naphthyl ring, exerts a spatial influence on the manifestation of 1H NMR signals, alongside other electronic effects. This study will offer NMR data, providing insights into the potential applications and analyses of naphthalene derivatives across various theoretical and practical domains. [ABSTRACT FROM AUTHOR]

A motivating class of compounds with interest in the research field of biological active metallopharmaceuticals for cancer treatment is based on organometallic complexes of Sn(IV), exhibiting advantages such as improved cellular uptake and body excretion, lower toxicity, and fewer side effects compared to platinum-based drugs. In this study, the mononuclear organotin coordination complex [(CH3)2SnCl2(mebta)2] was synthesized and characterized using vibrational spectroscopy (IR, Raman), 1H NMR, 13C{1H} NMR, and X-ray crystallography. Its antiproliferative properties were thoroughly assessed across an aggressive triple-negative human breast cancer cell line. Notably, comparative studies with precursor materials verified that the observed biological activity is intrinsic to the complex itself. This study highlights the compound's ability to induce cell fate by disrupting essential cellular functions, such as proliferation. By exploring the antiproliferative effects of organotin(IV) derivatives, we introduce a novel class of Sn complexes with 1-methylbenzotriazole (mebta), demonstrating significant potential as promising antitumor agents in the field of organotin compounds. [ABSTRACT FROM AUTHOR]

The success of platinum-based chemotherapeutic drugs for clinical cancer treatments has inspired tremendous research efforts on developing new metallic anticancer agents with improved cytotoxic activity and reduced side effects. 2,2′;6′,2″-Terpyridine and its 4′-substituted derivatives have showed great potential as ligand compartments for designing new anticancer drug candidates involving base metals. In this work, we synthesized a series of cobalt and iron coordination compounds based on 4′-pyridyl-2,2′;6′,2″-terpyridine, including homoleptic complexes, a dinuclear bridged complex and 1- and 2-dimensional coordination polymers/networks. The polymorphism of two homoleptic CoII and FeII complexes has been described along with the structural characterization of a CoII coordination polymer and dinuclear FeIII complex by X-ray crystallography. These compounds were tested preliminarily as precatalysts for the regioselective hydrosilylation of styrene. Their cytotoxic activities against two human breast cancer cell lines (MCF-7 and MDA-MB 468) and a normal breast epithelial cell line (MCF-10A) were investigated in order to observe the best-performing drug candidates. [ABSTRACT FROM AUTHOR]

Guided by the idea that the presence of a heterocyclic aromatic core and tyramine moiety, under the umbrella of a single molecular scaffold could bring interesting biological properties, herein we present synthesis, characterization, with two crystal structures reported, and biological evaluation of some tyramine derivates. Cytotoxic and antimigratory potential was addressed by using a colorectal cancer cell line as a model system. Although possessing no cytotoxic effects, two compounds have shown strong antimigratory potential in low doses, with no effect on healthy MRC‐5 cells. Evaluation of their antimicrobial activities suggested prominent antimicrobial activity, where Compound 4 outperformed streptomycin against Escherichia coli and Proteus mirabilis. Hormone‐dependent types of cancer, such as prostate, ovary, and breast, are highly dependent on human sex hormone–binding globulin (SHBG) blood levels. A molecular docking study has shown that 1 has high affinity to bind and therefore compete with natural steroids for the SHBG steroid‐binding site. DNA‐binding study have shown that 4 interacts with CT‐DNA in a groove‐binding mode. In silico ADME/T study revealed that all compounds have suitable physicochemical properties for oral bioavailability and druglikeness, while toxicity tests for 1, 4, and 6 suggested potential for mutagenicity (4, 6), hepatotoxicity (6), and skin sensation (1). [ABSTRACT FROM AUTHOR]

The conservation of the main protease in viral genomes, combined with the absence of a homologous protease in humans, makes this enzyme family an ideal target for developing broad-spectrum antiviral drugs with minimized host toxicity. GC-376, a peptidomimetic 3CL protease inhibitor, has shown significant efficacy against coronaviruses. Recently, a GC-376-based PROTAC was developed to target and induce the proteasome-mediated degradation of the dimeric SARS-CoV-2 3CLPro protein. Extending this approach, the current study investigates the application of the GC-376 PROTAC to the 3CPro protease of enteroviruses, specifically characterizing its interaction with CVB3 3CPro through X-ray crystallography, NMR (Nuclear Magnetic Resonance) and biochemical techniques. The crystal structure of CVB3 3CPro bound to the GC-376 PROTAC precursor was obtained at 1.9 Å resolution. The crystallographic data show that there are some changes between the binding of CVB3 3CPro and SARS-CoV-2 3CLPro, but the overall similarity is strong (RMSD on C-alpha 0.3 Å). The most notable variation is the orientation of the benzyloxycarbonyl group of GC-376 with the S4 subsite of the proteases. NMR backbone assignment of CVB3 3CPro bound and unbound to the GC-376 PROTAC precursor (80% and 97%, respectively) was obtained. This information complemented the investigation, by NMR, of the interaction of CVB3 3CPro with the GC-376 PROTAC, and its precursor allows us to define that the GC-376 PROTAC binds to CVB3 3CPro in a mode very similar to that of the precursor. The NMR relaxation data indicate that a quench of dynamics of a large part of the protein backbone involving the substrate-binding site and surrounding regions occurs upon GC-376 PROTAC precursor binding. This suggests that the substrate cavity, by sampling different backbone conformations in the absence of the substrate, is able to select the suitable one necessary to covalently bind the substrate, this being the latter reaction, which is the fundamental step required to functionally activate the enzymatic reaction. The inhibition activity assay showed inhibition potency in the micromolar range for GC-376 PROTAC and its precursor. Overall, we can conclude that the GC-376 PROTAC fits well within the binding sites of both proteases, demonstrating its potential as a broad-spectrum antiviral agent. [ABSTRACT FROM AUTHOR]

The voltage-dependent anion channel (VDAC) is a crucial mitochondrial protein that facilitates ion and metabolite exchange between mitochondria and the cytosol. Initially characterized over three decades ago, the structure of VDAC-1 was resolved in 2008, revealing a novel β-barrel protein architecture. This study presents the first room-temperature crystal structure of mouse VDAC-1 (mVDAC-1), which is a significant step toward understanding the channel's gating mechanism. The new structure, obtained at a 3.3 Å resolution, demonstrates notable differences from the previously determined cryogenic structure, particularly in the loop regions, which may be critical for the transition between the 'open' and 'closed' states of VDAC-1. Comparative analysis of the root-mean-square deviation (R.M.S.D.) and B-factors between the cryogenic and room-temperature structures suggests that these conformational differences, although subtle, are important for VDAC's functional transitions. The application of electric field-stimulated X-ray crystallography (EF-X) is proposed as a future direction to resolve the 'closed' state of VDAC-1 by inducing voltage-driven conformational changes in order to elucidate the dynamic gating mechanism of VDAC-1. Our findings have profound implications for understanding the molecular basis of VDAC's role in mitochondrial function and its regulation under physiological conditions. [ABSTRACT FROM AUTHOR]

Structural DNA nanotechnology is an emerging field and is expected to be used for various applications in materials science. In this study, we designed a DNA tensegrity triangle to accommodate the bipyridine complexes with metal ions (Ni2+ and Fe2+) at the center of the space within the triangle. A metal–bipyridine-incorporated DNA tensegrity triangle was crystalized, and the presence of metals within it was confirmed through X-ray crystal structure analysis. A signal of the anomalous dispersion effect derived from metal was observed in the center of the DNA triangle. [ABSTRACT FROM AUTHOR]

The antimicrobial properties of a series of anionic bis(carbene) silver complexes Na3[Ag(NHCR)2] were investigated (2a–2g and 2c′, where NHCR is a 2,2′-(imidazol-2-ylidene)dicarboxylate-type N-heterocyclic carbene). The complexes were synthesized by the interaction of imidazolium dicarboxylate compounds with silver oxide in the presence of aqueous sodium hydroxide. Complexes 2f,g were characterized analytically and spectroscopically, and the ligand precursor 1f and complexes 2c and 2g were structurally identified by X-ray diffraction methods. The anions of 2c and 2g, [Ag(NHCR)2]3−, showed a typical linear disposition of Ccarbene-Ag-Ccarbene atoms and an uncommonly eclipsed conformation of carbene ligands. The antimicrobial properties of complexes 2a–g, which contains chiral (2b–2e and 2c′) and non-chiral derivatives (2a,f,g), were evaluated against Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa, and a Gram-positive bacterium, Staphylococcus aureus. From the observed values of the minimal inhibitory concentration and minimal bactericidal concentration, complexes 2a and 2b showed the best antimicrobial activity against all strains. An interesting chirality–antimicrobial relationship was found, and eutomer 2c′ showed better activity than its enantiomer 2c against the three bacteria. Furthermore, these complexes were investigated experimentally and theoretically by 109Ag nuclear magnetic resonance, and the electronic and steric characteristics of the dianionic carbene ligands were also examined. [ABSTRACT FROM AUTHOR]

Fluorescent markers play important roles in spectroscopic and microscopic research techniques and are broadly used in basic and applied sciences. We have obtained markers with fluorescent properties, two etheno derivatives of 2-aminopurine, as follows: 1,N2-etheno-2-aminopurine (1,N2-ε2APu, I) and N2,3-etheno-2-aminopurine (N2,3-ε2APu, II). In the present paper, we investigate their interaction with two key enzymes of purine metabolism, purine nucleoside phosphorylase (PNP), and xanthine oxidase (XO), using diffraction of X-rays on protein crystals, isothermal titration calorimetry, and fluorescence spectroscopy. Crystals were obtained and structures were solved for WT PNP and D204N-PNP mutant in a complex with N2,3-ε2APu (II). In the case of WT PNP—1,N2-ε2APu (I) complex, the electron density corresponding to the ligand could not be identified in the active site. Small electron density bobbles may indicate that the ligand binds to the active site of a small number of molecules. On the basis of spectroscopic studies in solution, we found that, in contrast to PNP, 1,N2-ε2APu (I) is the ligand with better affinity to XO. Enzymatic oxidation of (I) leads to a marked increase in fluorescence near 400 nm. Hence, we have developed a new method to determine XO activity in biological material, particularly suitable for milk analysis. [ABSTRACT FROM AUTHOR]

During the automatic processing of crystallographic diffraction experiments, beamstop shadows are often unaccounted for or only partially masked. As a result of this, outlier reflection intensities are integrated, which is a known issue. Traditional statistical diagnostics have only limited effectiveness in identifying these outliers, here termed Not‐Excluded‐unMasked‐Outliers (NEMOs). The diagnostic tool AUSPEX allows visual inspection of NEMOs, where they form a typical pattern: clusters at the low‐resolution end of the AUSPEX plots of intensities or amplitudes versus resolution. To automate NEMO detection, a new algorithm was developed by combining data statistics with a density‐based clustering method. This approach demonstrates a promising performance in detecting NEMOs in merged data sets without disrupting existing data‐reduction pipelines. Re‐refinement results indicate that excluding the identified NEMOs can effectively enhance the quality of subsequent structure‐determination steps. This method offers a prospective automated means to assess the efficacy of a beamstop mask, as well as highlighting the potential of modern pattern‐recognition techniques for automating outlier exclusion during data processing, facilitating future adaptation to evolving experimental strategies. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Porphyromonas gingivalis is a major pathogenic oral bacterium that is responsible for periodontal disease. It is linked to chronic periodontitis, gingivitis and aggressive periodontitis. P. gingivalis exerts its pathogenic effects through mechanisms such as immune evasion and tissue destruction, primarily by secreting various factors, including cysteine proteases such as gingipain K (Kgp), gingipain R (RgpA and RgpB) and PrtH (UniProtKB ID P46071). Virulence proteins comprise multiple domains, including the pro‐peptide region, catalytic domain, K domain, R domain and DUF2436 domain. While there is a growing database of knowledge on virulence proteins and domains, there was no prior evidence or information regarding the structure and biological function of the well conserved DUF2436 domain. In this study, the DUF2436 domain of PrtH from P. gingivalis (PgDUF2436) was determined at 2.21 Å resolution, revealing a noncanonical β‐jelly‐roll sandwich topology with two antiparallel β‐sheets and one short α‐helix. Although the structure of PgDUF2436 was determined by the molecular‐replacement method using an AlphaFold model structure as a template, there were significant differences in the positions of β1 between the AlphaFold model and the experimentally determined PgDUF2436 structure. The Basic Local Alignment Search Tool sequence‐similarity search program showed no sequentially similar proteins in the Protein Data Bank. However, DaliLite search results using structure‐based alignment revealed that the PgDUF2436 structure has structural similarity Z‐scores of 5.9–5.4 with the C‐terminal domain of AlgF, the D4 domain of cytolysin, IglE and the extracellular domain structure of PepT2. This study has elucidated the structure of the DUF2436 domain for the first time and a comparative analysis with similar structures has been performed. [ABSTRACT FROM AUTHOR]