Journal of Biological Inorganic Chemistry最新文献

The last decade has seen a lot of interest in classical organometallic catalysis performed intracellularly, or at least under biological conditions (37 °C, air, water). One such classical reaction is olefin metathesis (OM), which is extremely common in preparative organic chemistry, however under non-biological conditions (esp. organic solvents). For in vivo applications, both prokaryotic and eukaryotic applications would require the passage of the OM catalyst through the cell membrane, which already contains unsaturated olefins and thus potential substrates for OM. This work focuses on the question whether OM is catalysed in membranes containing unsaturated phospholipids. Initial experiments with the 2nd generation Hoveyda-Grubbs catalyst (HGII) showed self-metathesis of the membrane-forming phospholipids POPE, POPC, and POPG, while approximating biological conditions in solution (37 °C, air, neutral pH) with substrate conversions up to 58%. Subsequent experiments with DOPC in large unilamellar vesicles (LUVs) and in solution included a PEGylated (featuring increased water solubility) and a palmitoylated (with increased membrane mobility) HGII derivative. Both were successfully synthesised beforehand and comprehensively characterized. Membrane localisation of the catalysts was evaluated via size exclusion chromatography (SEC) followed by ICP-MS. Product analysis of OM was carried out after one hour and 24 h, respectively, via NMR, TLC, and LC-MS. The intra- and intermolecular OM products of DOPC were identified. First, substrate conversion was significantly diminished in the vesicles, however, the equilibrium state in vesicles was reached in much less time compared to solution (1 h in vesicles vs. 24 h in solution). To our surprise, a distinct difference in product selectivity between OM in solution and in vesicles was observed in that the intramolecular OM is much favoured in solution, while in LUVs, the main products are the result of intermolecular OM reactions. Our results prove that OM is readily catalysed in vesicles and indicate that the milieu of the lipid bilayer has a major impact on the product selectivity, reaction time and substrate conversion. While previous work has focused on the cytotoxicity of Hoveyda-Grubbs catalysts and their interaction with biomolecules, our present work provides valuable insights on what will happen when an OM catalyst like HGII is exposed to membranes in organisms. Moreover, using the results from this work, it may be possible to selectively modify membrane properties, and thereby cellular responsiveness to outside stress, by olefin metathesis in the future.

Protein proclivity to aggregation leads to the formation of insoluble aggregates which slowly build up over the years within blood and different tissues. In humans, the content of these detergent resistant aggregates has been found increased in elder subjects compared to younger ones. The aggregation process is driven not solely by the presence of prone to aggregation proteins but by pro-aggregating agents such as Cu(II) which promotes the aggregation of IgG among others. Cu(II) and Zn(II) induce IgG reversible aggregates. However, co-incubation of Cu(II) with hydrogen peroxide (H₂O₂) renders the aggregate irreversibly insoluble upon metal removal which also becomes detergent resistant. Conversely, in the case of Zn(II)-induced aggregates, the addition of H₂O₂ in concentrations as high as 3 mM does not yield a detergent resistant aggregate. Contrarily to Cu(II), Zn(II) is redox inactive and unable to react with H₂O₂. Notably, the incubation of IgG with Cu(II) and low concentrations of H₂O₂ (50 µM) leads to the formation of soluble aggregates which can be detected by SDS-PAGE. Boiling allows partial dis-aggregation of the SDS-resistant aggregates and the migration pattern of boiled IgG oxidized in vitro by Cu(II)/H₂O₂ is similar to that observed in the aggregates from human sera. The presence of IgG was confirmed by western blot in SDS-resistant aggregates found in elder and young sera. Therefore, we speculate that Cu(II) reaction with H₂O₂ may contribute to IgG-enriched circulating protein aggregates and their clinical relevance should be further explored as they could resemble circulating immune-complexes which possess several pro-inflammatory features.

Mechanism of action (MoA) studies on the cytotoxic thiolato-bridged triosmium carbonyl clusters Os₃(CO)₁₀(μ-H)(μ-SR) (2) indicates that their cytotoxicity is associated with increased reactive oxygen species (ROS) generation, G₂/M cell cycle arrest, and subsequent apoptosis. Cellular uptake is a key factor, with an increased reactivity of the cluster with serum leading to reduced available concentrations in the medium thereby diminishing its anti-proliferative effect. Reactivity studies reveal that biomolecular interactions occur predominantly at the triosmium core, with a preference for amine-containing species.

Nitric oxide synthase (NOS) is an enzyme responsible for the production of nitric oxide in living organisms. Structurally, it is a homodimer composed of multiple domains connected by random coil tethers. The resulting structural flexibility, along with the diverse conformational states it enables, is essential for NOS function and remains an active area of investigations. Here, we studied the docking interactions between the reductase domains of NOS subunits. To probe these interactions, a nitroxide-based bifunctional spin label was attached to each T34C/S38C calmodulin (CaM) molecule bound to the CaM-binding region of the tether, which connects the oxygenase and flavin mononucleotide (FMN) domains in each subunit of the homodimeric oxygenase/FMN (oxyFMN) construct of rat neuronal NOS (nNOS). The magnetic dipole interaction between the spin labels was detected by 2 + 1 electron spin echo (ESE) methods. The experimental 2 + 1 ESE traces were interpreted using the Monte Carlo calculations of NOS conformational distributions. The results unequivocally show that at the estimated effective temperature of the frozen conformational distribution, T_ef ≈ 200 K, a large proportion of the oxyFMN proteins (~ 55%) adopt a clamp-shaped conformation in which the FMN domains of different NOS subunits dock with each other. The stabilization energy of this docking complex (i.e., docking energy) was estimated in the model of isotropic interaction as - 7.2kT_ef ≈ - 2.9 kcal/mol. The identification of this clamp-shaped conformation suggests it as an intermediate structural state that may influence NOS catalytic efficiency by facilitating the FMN–heme interdomain electron transfer through constraining the conformational space accessible to the FMN domain as it approaches its docking positions at the heme domain.

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Non-canonical DNA structures play important roles in processing of the genetic code. Three-way (3WJ) and four-way (4WJ) junctions are dynamic, multi-stranded structures containing an open cavity at the centre. We have previously demonstrated that supramolecular dinuclear metallo-cylinders bind well inside 3WJ cavities, having an optimally complementary size and shape match, cationic charge to bind the DNA anion, as well as the ability to π‑stack with the branchpoint nucleobases. Herein, we show that a longer metallo-cylinder with a similar but extended central π surface binds to both 3WJ and 4WJ structures with good selectivity over double-stranded DNA. Experimental investigations, informed by molecular dynamics (MD) simulations, reveal that whilst this longer cylinder can bind 3WJs as the previously studied cylinders, the extended π surface of the cylinder now also facilitates 4WJ binding. The simulations capture two metastable 4WJ conformations –one resembling a 3WJ, and another where the extended length enables the cylinder to angle into and stabilise a rhombus-shaped 4WJ cavity. The ability to tune the structure of supramolecular assemblies is important for targeting different DNA structures with varying specificity, and in this work, we demonstrate the usefulness of overall length as a parameter for modulating DNA binding.

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A low-molecular-weight analog of the 1-aminocyclopropane carboxylic acid oxidase (ACCO), [Tp^MesFeACC], 1, where the (His₂Asp)iron(II) moiety is mimicked by a hydrotris(3-mesitylpyrazol-1-yl)borato iron(II) unit, to which the natural substrate aminocyclopropane carboxylate is coordinated, has been accessed and structurally characterized. It was found to react slowly with O₂ to yield the biological product ethylene. To create models of the intermediates proposed as part of the catalytic cycle of the ACCO 1 was treated with tBuOOH and mCPBA at low temperatures, which generated the respective Fe^IIIOOtBu and Fe^IV=O intermediates as shown by spectroscopic analysis. Studies on their behavior upon annealing reveal a non-biomimetic reactivity.

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Vimentin is a principal intermediate filament (IF) protein that is essential for maintaining cytoskeleton architecture and cellular mechanical integrity. Growing evidence is revealing that metal ions play critical roles in modulating the structure, assembly, and mechanics of vimentin IFs. Despite this, a detailed molecular-level understanding of vimentin–metal interactions and its functional consequences remains incomplete. This review summarizes the current knowledge of metal-induced effects on the structural and mechanical properties of vimentin and highlights how post-translational modifications and cytoskeletal dynamics may alter these metal–protein interactions. Advancing our understanding of the interplay between metal ions and vimentin IFs will enhance our comprehension of the complex mechanisms governing the versatile functions of vimentin and other IF proteins in health and disease.

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A novel ruthenium(II) compound, cis-[Ru(phen)₂(4-bzpy)(Cl)](PF₆) (complex I), where phen = 1,10-phenanthroline and 4-bzpy = 4-benzoylpyridine, was synthesized and fully characterized using electrochemical and spectroscopic methods, and its structure was determined by single-crystal X-ray diffraction. Density functional theory (DFT) and time-dependent DFT calculations were performed to shed light on the electronic structure and nature of the vibrational and electronic transitions. The photochemical behavior of complex I in aqueous solution showed that irradiation with blue light (453 nm) promoted the release of the coordinated 4-bzpy ligand originating cis-[Ru(phen)₂(H₂O)(Cl)]⁺ ion, as identified by NMR and electronic spectroscopy. Moreover, complex I exhibited a great ability to cleave DNA molecules upon blue light irradiation, which was associated with the production of reactive oxygen species (singlet oxygen and superoxide anion). In this study, we also investigated the antimicrobial activity of complex I along with a similar compound, cis-[Ru(bpy)₂(4-bzpy)(Cl)](PF₆) (complex II), their precursors [Ru(bpy)₂Cl₂] and [Ru(phen)₂Cl₂], and the free ligand 4-bzpy. These two ruthenium complexes I and II have a common auxiliary ligand 4-bzpy, but distinct chelating ligands (phenanthroline or 2,2′-bipyridine, bpy). Notably, both complexes showed promising antibacterial activity against Gram-positive bacterial strains of Staphylococcus aureus and Staphylococcus epidermidis. However, complex I showed a superior antibacterial effect compared with complex II, supporting the important role of the phen ligand, likely providing greater lipophilicity to this compound.

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Chamomile extract (CHA) was employed in the synthesis of silver nanoparticles (AgNPs(CHA)). These nanoparticles were incorporated into a hydroxyethyl methacrylate (pHEMA) matrix during polymerization at concentrations of 1 mg/mL (pHEMA@AgNPs(CHA)_1) and 2 mg/mL (pHEMA@AgNPs(CHA)_2). The resulting materials— AgNPs(CHA), pHEMA@AgNPs(CHA)_1, and pHEMA@AgNPs(CHA)_2 —were characterized using a plethora of analytical techniques, including Refractive Index (RI), X-ray fluorescence (XRF) spectroscopy, X-ray powder diffraction (XRPD), thermogravimetric-differential thermal analysis (TG–DTA), differential scanning calorimetry (DSC), ultraviolet–visible (UV–Vis) spectroscopy, and attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy in the solid state, as well as UV–Vis spectroscopy in solution. XRPD analysis revealed a crystallite size of 8.02 nm, while the diameter of the nanoparticles, estimated using the λmax of the Surface Plasmon Resonance (SPR) spectrum, was 37 nm. The antimicrobial properties of the materials were evaluated against Gram-negative bacteria, Pseudomonas aeruginosa and Escherichia coli, as well as Gram-positive bacteria, Staphylococcus epidermidis and Staphylococcus aureus. These pathogens are commonly implicated in microbial keratitis. Antimicrobial efficacy was assessed using Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), and Inhibitory Zones (IZs). The IZs for P. aeruginosa, E. coli, S. epidermidis, and S. aureus following incubation with paper discs soaked in 1 mg/mL of AgNPs(CHA) were 16.0 ± 0.6 mm, 11.1 ± 0.9 mm, 15.8 ± 1.2 mm, and 15.1 ± 0.2 mm, respectively. Bacterial viability after incubation with hydrogel discs of pHEMA@AgNPs(CHA)_1 showed reductions to 0.9 ± 0.2%, 100.0 ± 0.0%, 1.9 ± 1.0%, and 4.4 ± 1.5% for P. aeruginosa, E. coli, S. epidermidis, and S. aureus, respectively. Additionally, no toxic effects were observed on human corneal epithelial cells (HCECs). These findings suggest that pHEMA@AgNPs(CHA)_1 has potential as a promising candidate for developing non-infectious contact lenses.

Magnetic interactions between iron–sulfur (Fe/S) clusters and transition metal centers such as nickel, molybdenum, and copper play a central role in the function of key metalloenzymes. These interactions, which arise from electronic coupling, spin exchange, and spatial arrangement, directly influence redox behavior and catalytic efficiency. This review highlights three distinct complex enzymes—[NiFe] hydrogenases, mononuclear molybdenum-containing xanthine oxidase (XO) family, and [NiFe] and [MoCu] carbon monoxide dehydrogenases (CODHs)—as paradigms for understanding (Fe/S)-metal center interactions. In [NiFe] hydrogenases, (Fe/S) clusters serve as electron relays that magnetically interact with the catalytic [NiFe] active site. In XO-type enzymes, a mononuclear Mo center is functionally and magnetically coupled to nearby Fe/S clusters, modulating substrate reduction and electron transfer. Similarly, in CODHs, both [NiFe]—and [MoCu]-dependent variants exhibit strong magnetic communication between metal active sites and surrounding Fe/S clusters, crucial for CO₂/CO interconversion. Advanced spectroscopic approaches, particularly electron paramagnetic resonance (EPR) and related techniques, combined with theoretical modelling, have provided deep insights into the electronic structures and dynamic interactions within these metalloenzymes. Understanding these magnetic interactions not only sheds light on fundamental electron-transfer and enzymatic mechanisms but also guides the design of bioinspired catalysts and energy-conversion technologies.

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