Gating movements and ion permeation in HCN4 pacemaker channels.

The HCN1-4 channel family is responsible for the hyperpolarization-activated cation current If/Ih that controls automaticity in cardiac and neuronal pacemaker cells. We present cryoelectron microscopy (cryo-EM) structures of HCN4 in the presence or absence of bound cAMP, displaying the pore domain in closed and open conformations. Analysis of cAMP-bound and -unbound structures sheds light on how ligand-induced transitions in the channel cytosolic portion mediate the effect of cAMP on channel gating and highlights the regulatory role of a Mg2+ coordination site formed between the C-linker and the S4-S5 linker. Comparison of open/closed pore states shows that the cytosolic gate opens through concerted movements of the S5 and S6 transmembrane helices. Furthermore, in combination with molecular dynamics analyses, the open pore structures provide insights into the mechanisms of K+/Na+ permeation. Our results contribute mechanistic understanding on HCN channel gating, cyclic nucleotide-dependent modulation, and ion permeation.

Protein lipoylation: mitochondria, cuproptosis, and beyond.

Protein lipoylation, a crucial post-translational modification (PTM), plays a pivotal role in mitochondrial function and emerges as a key player in cell death through cuproptosis. This novel copper-driven cell death pathway is activated by excessive copper ions binding to lipoylated mitochondrial proteins, disrupting energy production and causing lethal protein aggregation and cell death. The intricate relationship among protein lipoylation, cellular energy metabolism, and cuproptosis offers a promising avenue for regulating essential cellular functions. This review focuses on the mechanisms of lipoylation and its significant impact on cell metabolism and cuproptosis, emphasizing the key genes involved and their implications for human diseases. It offers valuable insights into targeting dysregulated cellular metabolism for therapeutic purposes.

Evolution of Inosine-Specific Endonuclease V from Bacterial DNase to Eukaryotic RNase.

Endonuclease V (EndoV) cleaves the second phosphodiester bond 3' to a deaminated adenosine (inosine). Although highly conserved, EndoV homologs change substrate preference from DNA in bacteria to RNA in eukaryotes. We have characterized EndoV from six different species and determined crystal structures of human EndoV and three EndoV homologs from bacteria to mouse in complex with inosine-containing DNA/RNA hybrid or double-stranded RNA (dsRNA). Inosine recognition is conserved, but changes in several connecting loops in eukaryotic EndoV confer recognition of 3 ribonucleotides upstream and 7 or 8 bp of dsRNA downstream of the cleavage site, and bacterial EndoV binds only 2 or 3 nt flanking the scissile phosphate. In addition to the two canonical metal ions in the active site, a third Mn2+ that coordinates the nucleophilic water appears necessary for product formation. Comparison of EndoV with its homologs RNase H1 and Argonaute reveals the principles by which these enzymes recognize RNA versus DNA.

Insertion of CO2 in metal ion-doped two-dimensional covalent organic frameworks.

Carbon capture is one of the essential low-carbon technologies required to achieve societal climate goals at the lowest cost. Covalent organic frameworks (COFs) are promising adsorbents for CO2 capture because of their well-defined porosity, large surface area, and high stability. Current COF-based CO2 capture is mainly based on a physisorption mechanism, exhibiting smooth and reversible sorption isotherms. In the present study, we report unusual CO2 sorption isotherms featuring one or more tunable hysteresis steps with metal ion (Fe3+, Cr3+, or In3+)-doped Schiff-base two-dimensional (2D) COFs (Py-1P, Py-TT, and Py-Py) as adsorbents. Synchrotron X-ray diffraction, spectroscopic and computational studies indicate that the sharp adsorption steps in the isotherm originate from the insertion of CO2 between the metal ion and the N atom of the imine bond on the inner pore surface of the COFs as the CO2 pressure reaches threshold values. As a result, the CO2 adsorption capacity of the ion-doped Py-1P COF is increased by 89.5% compared with that of the undoped Py-1P COF. This CO2 sorption mechanism provides an efficient and straightforward approach to enhancing the CO2 capture capacity of COF-based adsorbents, yielding insights into developing chemistry for CO2 capture and conversion.

Exploring the molecular mechanism of coloration differences in two Meconopsis wilsonii subspecies: australis and orientalis.

Flower color diversity is a key taxonomic trait in Meconopsis species, enhancing their appeal as ornamental flowers. However, knowledge of the molecular mechanisms of flower color formation in Meconopsis species is still limited. M. wilsonii subsp. australis (Australis) and M. wilsonii subsp. orientalis (Orientalis) have a developmental stage presenting red-purple flowers, while Orientalis also presents blue coloration at the full-bloom period, making them an important model for exploring the mechanism of blue flower formation in M. wilsonii. In this study, we collected petals from Australis and Orientalis at different developmental stages to compare the coloration differences between the two species and detect the molecular mechanisms of blue color in Orientalis. We identified that cyanidin was the main anthocyanin in the flowers of both species, and the blue color in Orientalis primarily arises from anthocyanins (Cyanidin-3-O-sambubioside). RNA sequencing analysis was performed to detect the gene expression in the anthocyanin biosynthesis pathway, and the results suggested that gene regulation for anthocyanin biosynthesis may not be the direct reason for blue color formation in Orientalis. In addition, the growth solid of Orientalis was rich in Fe and Mg ions, and a large amount of Fe and Mg ions accumulated in the petals of Orientalis. Combined with the gene functional enrichment results, we found that the purple and red-purple colors of these two species were presented by different glycosylation levels of cyanidin, while the violet color of Orientalis might be the results of metalloanthocyanins by Fe and Mg ions, which also relieved the toxicity caused by the high content of Fe and Mg ions in its cells. The environmental adaptation-related genes were highly expressed of in both species, such as adaptation to desiccation, water deprivation, freezing, etc. Our results revealed the coloration differences between Australis and Orientalis and described the molecular mechanisms of blue coloration in Orientalis. The data in our analysis could enrich the genetic resources for M. wilsonii for further studies.

Identification of metal ion-binding sites in RNA structures using deep learning method.

Metal ion is an indispensable factor for the proper folding, structural stability and functioning of RNA molecules. However, it is very difficult for experimental methods to detect them in RNAs. With the increase of experimentally resolved RNA structures, it becomes possible to identify the metal ion-binding sites in RNA structures through in-silico methods. Here, we propose an approach called Metal3DRNA to identify the binding sites of the most common metal ions (Mg2+, Na+ and K+) in RNA structures by using a three-dimensional convolutional neural network model. The negative samples, screened out based on the analysis for binding surroundings of metal ions, are more like positive ones than the randomly selected ones, which are beneficial to a powerful predictor construction. The microenvironments of the spatial distributions of C, O, N and P atoms around a sample are extracted as features. Metal3DRNA shows a promising prediction power, generally surpassing the state-of-the-art methods FEATURE and MetalionRNA. Finally, utilizing the visualization method, we inspect the contributions of nucleotide atoms to the classification in several cases, which provides a visualization that helps to comprehend the model. The method will be helpful for RNA structure prediction and dynamics simulation study. Availability and implementation: The source code is available at https://github.com/ChunhuaLiLab/Metal3DRNA.

Bodipy-based quinoline derivative as a highly Hg2+-selective fluorescent chemosensor and its potential applications.

A highly efficient sensor has been successfully developed using quinoline-based BODIPY compounds (8-quinoline-4,4-difluoro-4-boro-3a, 4a-diazaindacene (C1) and 7-hydroxy-8-quinoline-4,4-difluoro-4-boro-3a, 4a-diazindacene (C2) to detect Hg2+ ions. The sensor C1 exhibits remarkable selectivity in detecting Hg2+ with a limit of detection 3.06 × 10-8 mol/L. The developed chemical sensors have shown stability, cost-effectiveness, ease of preparation, and remarkable selectivity towards Hg2+ ions compared to other commonly occurring metal ions. The total recovery of the sensor C1 can be achieved by using a 0.1 mol/L solution of KI. The proposed sensor C1 has been applied to determine Hg2+ in tap and distilled water, yielding excellent results. In addition, the binding mode of C1-Hg2+ and C2-Hg2+ complexes was a 1:1 ratio confirmed by mass spectra, Job's plot, and DFT study. Moreover, the sensor C1 successfully applied for the biological studies results in negligible cytotoxicity, which demonstrates it can be used to determine Hg2+ in HT22 cells.

Visualization of mutagenic nucleotide processing by Escherichia coli MutT, a Nudix hydrolase.

Escherichia coli MutT prevents mutations by hydrolyzing mutagenic 8-oxo-2'-deoxyguanosine 5'-triphosphate (8-oxo-dGTP) in the presence of Mg2+ or Mn2+ ions. MutT is one of the most studied enzymes in the nucleoside diphosphate-linked moiety X (Nudix) hydrolase superfamily, which is widely distributed in living organisms. However, the catalytic mechanisms of most Nudix hydrolases, including two- or three-metal-ion mechanisms, are still unclear because these mechanisms are proposed using the structures mimicking the reaction states, such as substrate analog complexes. Here, we visualized the hydrolytic reaction process of MutT by time-resolved X-ray crystallography using a biological substrate, 8-oxo-dGTP, and an active metal ion, Mn2+. The reaction was initiated by soaking MutT crystals in a MnCl2 solution and stopped by freezing the crystals at various time points. In total, five types of intermediate structures were refined by investigating the time course of the electron densities in the active site as well as the anomalous signal intensities of Mn2+ ions. The structures and electron densities show that three Mn2+ ions bind to the Nudix motif of MutT and align the substrate 8-oxo-dGTP for catalysis. Accompanied by the coordination of the three Mn2+ ions, a water molecule, bound to a catalytic base, forms a binuclear Mn2+ center for nucleophilic substitution at the ß-phosphorus of 8-oxo-dGTP. The reaction condition using Mg2+ also captured a structure in complex with three Mg2+ ions. This study provides the structural details essential for understanding the three-metal-ion mechanism of Nudix hydrolases and proposes that some of the Nudix hydrolases share this mechanism.

The battle for silver binding: How the interplay between the SilE, SilF, and SilB proteins contributes to the silver efflux pump mechanism.

The resistance of gram-negative bacteria to silver ions is mediated by a silver efflux pump, which mainly relies on a tripartite efflux complex SilCBA, a metallochaperone SilF and an intrinsically disordered protein SilE. However, the precise mechanism by which silver ions are extruded from the cell and the different roles of SilB, SilF, and SilE remain poorly understood. To address these questions, we employed nuclear magnetic resonance and mass spectrometry to investigate the interplay between these proteins. We first solved the solution structures of SilF in its free and Ag+-bound forms, and we demonstrated that SilB exhibits two silver binding sites in its N and C termini. Conversely to the homologous Cus system, we determined that SilF and SilB interact without the presence of silver ions and that the rate of silver dissociation is eight times faster when SilF is bound to SilB, indicating the formation of a SilF-Ag-SilB intermediate complex. Finally, we have shown that SilE does not bind to either SilF or SilB, regardless of the presence or absence of silver ions, further corroborating that it merely acts as a regulator that prevents the cell from being overloaded with silver. Collectively, we have provided further insights into protein interactions within the sil system that contribute to bacterial resistance to silver ions.

Nucleotide addition and cleavage by RNA polymerase II: Coordination of two catalytic reactions using a single active site.

RNA polymerase II (Pol II) incorporates complementary ribonucleotides into the growing RNA chain one at a time via the nucleotide addition cycle. The nucleotide addition cycle, however, is prone to misincorporation of noncomplementary nucleotides. Thus, to ensure transcriptional fidelity, Pol II backtracks and then cleaves the misincorporated nucleotides. These two reverse reactions, nucleotide addition and cleavage, are catalyzed in the same active site of Pol II, which is different from DNA polymerases or other endonucleases. Recently, substantial progress has been made to understand how Pol II effectively performs its dual role in the same active site. Our review highlights these recent studies and provides an overall model of the catalytic mechanisms of Pol II. In particular, RNA extension follows the two-metal-ion mechanism, and several Pol II residues play important roles to facilitate the catalysis. In sharp contrast, the cleavage reaction is independent of any Pol II residues. Interestingly, Pol II relies on its residues to recognize the misincorporated nucleotides during the backtracking process, prior to cleavage. In this way, Pol II efficiently compartmentalizes its two distinct catalytic functions using the same active site. Lastly, we also discuss a new perspective on the potential third Mg2+ in the nucleotide addition and intrinsic cleavage reactions.

Lanpepsy is a novel lanthanide-binding protein involved in the lanthanide response of the obligate methylotroph Methylobacillus flagellatus.

Lanthanides were recently discovered as metals required in the active site of certain methanol dehydrogenases. Since then, the characterization of the lanthanome, that is, proteins involved in sensing, uptake, and utilization of lanthanides, has become an active field of research. Initial exploration of the response to lanthanides in methylotrophs has revealed that the lanthanome is not conserved and that multiple mechanisms for lanthanide utilization must exist. Here, we investigated the lanthanome in the obligate model methylotroph Methylobacillus flagellatus. We used a proteomic approach to analyze differentially regulated proteins in the presence of lanthanum. While multiple known proteins showed induction upon growth in the presence of lanthanum (Xox proteins, TonB-dependent receptor), we also identified several novel proteins not previously associated with lanthanide utilization. Among these was Mfla_0908, a periplasmic 19 kDa protein without functional annotation. The protein comprises two characteristic PepSY domains, which is why we termed the protein lanpepsy (LanP). Based on bioinformatic analysis, we speculated that LanP could be involved in lanthanide binding. Using dye competition assays, quantification of protein-bound lanthanides by inductively coupled plasma mass spectrometry, as well as isothermal titration calorimetry, we demonstrated the presence of multiple lanthanide binding sites that showed selectivity over the chemically similar calcium ion. LanP thus represents the first member of the PepSY family that binds lanthanides. Although the physiological role of LanP is still unclear, its identification is of interest for applications toward the sustainable purification and separation of rare-earth elements.

The adaptability of the ion-binding site by the Ag(I)/Cu(I) periplasmic chaperone SilF.

The periplasmic chaperone SilF has been identified as part of an Ag(I) detoxification system in Gram-negative bacteria. Sil proteins also bind Cu(I) but with reported weaker affinity, therefore leading to the designation of a specific detoxification system for Ag(I). Using isothermal titration calorimetry, we show that binding of both ions is not only tighter than previously thought but of very similar affinities. We investigated the structural origins of ion binding using molecular dynamics and QM/MM simulations underpinned by structural and biophysical experiments. The results of this analysis showed that the binding site adapts to accommodate either ion, with key interactions with the solvent in the case of Cu(I). The implications of this are that Gram-negative bacteria do not appear to have evolved a specific Ag(I) efflux system but take advantage of the existing Cu(I) detoxification system. Therefore, there are consequences for how we define a particular metal resistance mechanism and understand its evolution in the environment.

Repair of topoisomerase 1-induced DNA damage by tyrosyl-DNA phosphodiesterase 2 (TDP2) is dependent on its magnesium binding.

Topoisomerases are enzymes that relax DNA supercoiling during replication and transcription. Camptothecin, a topoisomerase 1 (TOP1) inhibitor, and its analogs trap TOP1 at the 3'-end of DNA as a DNA-bound intermediate, resulting in DNA damage that can kill cells. Drugs with this mechanism of action are widely used to treat cancers. It has previously been shown that tyrosyl-DNA phosphodiesterase 1 (TDP1) repairs TOP1-induced DNA damage generated by camptothecin. In addition, tyrosyl-DNA phosphodiesterase 2 (TDP2) plays critical roles in repairing topoisomerase 2 (TOP2)-induced DNA damage at the 5'-end of DNA and in promoting the repair of TOP1-induced DNA damage in the absence of TDP1. However, the catalytic mechanism by which TDP2 processes TOP1-induced DNA damage has not been elucidated. In this study, we found that a similar catalytic mechanism underlies the repair of TOP1- and TOP2-induced DNA damage by TDP2, with Mg2+-TDP2 binding playing a role in both repair mechanisms. We show chain-terminating nucleoside analogs are incorporated into DNA at the 3'-end and abort DNA replication to kill cells. Furthermore, we found that Mg2+-TDP2 binding also contributes to the repair of incorporated chain-terminating nucleoside analogs. Overall, these findings reveal the role played by Mg2+-TDP2 binding in the repair of both 3'- and 5'-blocking DNA damage.

Recent Advances in Pristine Iron Triad Metal-Organic Framework Cathodes for Alkali Metal-Ion Batteries.

Pristine iron triad metal-organic frameworks (MOFs), i.e., Fe-MOFs, Co-MOFs, Ni-MOFs, and heterometallic iron triad MOFs, are utilized as versatile and promising cathodes for alkali metal-ion batteries, owing to their distinctive structure characteristics, including modifiable and designable composition, multi-electron redox-active sites, exceptional porosity, and stable construction facilitating rapid ion diffusion. Notably, pristine iron triad MOFs cathodes have recently achieved significant milestones in electrochemical energy storage due to their exceptional electrochemical properties. Here, the recent advances in pristine iron triad MOFs cathodes for alkali metal-ion batteries are summarized. The redox reaction mechanisms and essential strategies to boost the electrochemical behaviors in associated electrochemical energy storage devices are also explored. Furthermore, insights into the future prospects related to pristine iron triad MOFs cathodes for lithium-ion, sodium-ion, and potassium-ion batteries are also delivered.

High-Performance Bioinspired Microspheres for Boosting Dental Adhesion.

Dental adhesives are widely used in daily practice for minimally invasive restorative dentistry but suffer from bond degradation and biofilm attack. Bio-inspired by marine mussels having excellent surface-adhesion capability and high chemical affinity of polydopamine (PDA) to metal ions, herein, experimental zinc (Zn)-containing polydopamine-based adhesive formulation, further being referred to as "Zn-PDA@SiO2"-incorporated adhesive is proposed as a novel dental adhesive. Different Zn contents (5 and 10 mm) of Zn-PDA@SiO2 are prepared. Considering the synergistic effect of Zn and PDA, Zn-PDA@SiO2 not only presents excellent antibacterial potential and notably inhibits enzymatic activity (soluble and matrix-bound proteases), but also exhibits superior biocompatibility and biosafety in vitro/vivo. The long-term bond stability is substantially improved by adding 5 wt% 5 mm Zn-PDA@SiO2 to the primer. The aged bond strength of the experimentally formulated dental adhesives applied in self-etch (SE) bonding mode is 1.9 times higher than that of the SE gold-standard adhesive. Molecular dynamics calculations indicate the stable formation of covalent bonds, Zn-assisted coordinative bonds, and hydrogen bonds between PDA and collagen. Overall, this bioinspired dental adhesive provides an avenue technology for innovative biomedical applications and has already revealed promising perspectives for dental restorative dentistry.

Interlayer Entropy Engineering Inducing the Symmetry-Broken Layered Oxide Cathodes to Activate Reversible High-Voltage Redox Reaction.

The as-reported doping entropy engineering of electrode materials that are usually realized by the sharing of multiple metal elements with the metal element from the lattice body, potentially has three shortages of stringent synthesis conditions, large active element loss, and serious lattice distortion. Herein, an interlayer entropy engineering of layered oxide cathodes is proposed, where the multiple metal ions are simultaneously intercalated into the same interlayer sites, thus avoiding the three shortages. Concretely, a novel interlayer medium-entropy V2O5 ((MnCoNiMgZn)0.26V2O5∙0.84H2O) is successfully constructed by a one-step hydrothermal method. The interlayer medium-entropy effect is revealed to be that five metal ions pre-intercalation induces the local symmetry-broken [VO6] octahedra in bilayer V2O5, thus activating the reversible high-voltage redox reaction, inhibiting the layer slip and following phase transformation by its pinning effect, and enhancing the charge transfer kinetics. As a result, the medium-entropy cathode realizes the trade-off between specific capacity and structural stability with a discharge capacity of 152 mAh g-1 at 0.1 A g-1 after 100 cycles, and a capacity retention rate of 98.7% at 0.5 A g-1 after 150 cycles for Li+ storage. This engineering provides a new guideline for the rational design of high-performance layered oxide cathodes.

Multifunctional Antimonene-Silver Nanocomposites for Ultra-Multi-Mode and Multi-Analyte Sensing, Parallel and Batch Logic Computing, Long-Text Information Protection.

To simulate life's emergent functions, mining the multiple sensing capabilities of nanosystems, and digitizing networks of transduction signals and molecular interactions, is an ongoing endeavor. Here, multifunctional antimonene-silver nanocomposites (AM-Ag NCs) are synthesized facilely and fused for molecular sensing and digitization applications (including ultra-multi-mode and multi-analyte sensing, parallel and batch logic computing, long-text information protection). By mixing surfactant, AM, Ag+ and Sodium borohydride (NaBH4) at room temperature for 5 min, the resulting NCs are comprised of Ag nanoparticles scattered within AM nanosheets and protected by the surfactant. Interestingly, AM-Ag NCs exhibit ultra-multi-mode sensing ability for multiplex metal ions (Hg2+, Fe3+, or Al3+), which significantly improved selectivity (≈2 times) and sensitivity (≈400 times) when analyzing the combined channels. Moreover, multiple sensing capabilities of AM-Ag NCs enable diverse batch and parallel molecular logic computations (including advanced cascaded logic circuits). Ultra-multi-mode selective patterns of AM-Ag NCs to 18 kinds of metal ions can be converted into a series of binary strings by setting the thresholds, and realized high-density, long-text information protection for the first time. This study provides new ideas and paradigms for the preparation and multi-purpose application of 2D nanocomposites, but also offers new directions for the fusion of molecular sensing and informatization.

Alignment-free metal ion-binding site prediction from protein sequence through pretrained language model and multi-task learning.

More than one-third of the proteins contain metal ions in the Protein Data Bank. Correct identification of metal ion-binding residues is important for understanding protein functions and designing novel drugs. Due to the small size and high versatility of metal ions, it remains challenging to computationally predict their binding sites from protein sequence. Existing sequence-based methods are of low accuracy due to the lack of structural information, and time-consuming owing to the usage of multi-sequence alignment. Here, we propose LMetalSite, an alignment-free sequence-based predictor for binding sites of the four most frequently seen metal ions in BioLiP (Zn2+, Ca2+, Mg2+ and Mn2+). LMetalSite leverages the pretrained language model to rapidly generate informative sequence representations and employs transformer to capture long-range dependencies. Multi-task learning is adopted to compensate for the scarcity of training data and capture the intrinsic similarities between different metal ions. LMetalSite was shown to surpass state-of-the-art structure-based methods by more than 19.7, 14.4, 36.8 and 12.6% in area under the precision recall on the four independent tests, respectively. Further analyses indicated that the self-attention modules are effective to learn the structural contexts of residues from protein sequence. We provide the data sets, source codes and trained models of LMetalSite at https://github.com/biomed-AI/LMetalSite.

GASS-Metal: identifying metal-binding sites on protein structures using genetic algorithms.

Metals are present in >30% of proteins found in nature and assist them to perform important biological functions, including storage, transport, signal transduction and enzymatic activity. Traditional and experimental techniques for metal-binding site prediction are usually costly and time-consuming, making computational tools that can assist in these predictions of significant importance. Here we present Genetic Active Site Search (GASS)-Metal, a new method for protein metal-binding site prediction. The method relies on a parallel genetic algorithm to find candidate metal-binding sites that are structurally similar to curated templates from M-CSA and MetalPDB. GASS-Metal was thoroughly validated using homologous proteins and conservative mutations of residues, showing a robust performance. The ability of GASS-Metal to identify metal-binding sites was also compared with state-of-the-art methods, outperforming similar methods and achieving an MCC of up to 0.57 and detecting up to 96.1% of the sites correctly. GASS-Metal is freely available at https://gassmetal.unifei.edu.br. The GASS-Metal source code is available at https://github.com/sandroizidoro/gassmetal-local.

Water-Based Synthesis of ß-Sheet-Like Supramolecular Metallohydrogel Organized by Using a Native Ultrashort Peptide Sequence.

Designing supramolecular hydrogels using short peptides is challenging. To control self-assembly, a certain amount of organic solvent is typically added to the system, or the short peptide is modified with a functional group that is hydrophobic, hydrophilic, or highly coordinative. We discovered that l-His-l-Ile-l-Thr (HIT), a very short unmodified "native" tripeptide, selectively responds to Cu2+ ions in pure water to form a transparent supramolecular metallohydrogel. Circular dichroism analysis revealed that Cu2+ ions, but no other metal species, caused HIT to change from a random-coil-like to a ß-sheet-like structure. Other spectroscopic methods were used to characterize the properties of the supramolecular metallohydrogel. These results are expected to facilitate the development of native short peptides as advanced functional biomaterials.