The protective effect of ginsenoside Rg1 against sepsis-induced lung injury through PI3K-Akt pathway: insights from molecular dynamics simulation and experimental validation.

Sepsis-induced acute lung injury (SALI) poses a significant threat with high incidence and mortality rates. Ginsenoside Rg1 (GRg1), derived from Ginseng in traditional Chinese medicine, has been found to reduce inflammation and protect lung epithelial cells against tissue damage. However, the specific roles and mechanisms by which GRg1 mitigates SALI have yet to be fully elucidated. In this context, we employed a relevant SALI mouse model, alongside network pharmacology, molecular docking, and molecular dynamics simulation to pinpoint GRg1's action targets, complemented by in vitro assays to explore the underlying mechanisms. Our research shows that GRg1 alleviates CLP-induced SALI, decreasing lung tissue damage and levels of serum proinflammatory factor IL-6, TNF-α, and IL-1ß, also enhancing the survival rate of CLP mice. A total of 116 common targets between GRg1 and ALI, with specific core targets including AKT1, VEGFA, SRC, IGF1, ESR1, STAT3, and ALB. Further in vitro experiments assessed GRg1's intervention effects on MLE-12 cells exposed to LPS, with qRT-PCR analysis and molecular dynamics simulations confirming AKT1 as the key target with the favorable binding activity for GRg1. Western blot results indicated that GRg1 increased the Bcl-2/Bax protein expression ratio to reduce apoptosis and decreased the high expression of cleaved caspase-3 in LPS-induced MLE-12 cells. More results showed significant increases in the phosphorylation of PI3K and AKT1. Flow cytometric analysis using PI and Annexin-V assays further verified that GRg1 decreased the apoptosis rate in LPS-stimulated MLE-12 cells (from 14.85 to 6.54%, p < 0.05). The employment of the AKT1 inhibitor LY294002 confirmed these trends, indicating that AKT1's inhibition negates GRg1's protective effects on LPS-stimulated MLE-12 cells. In conclusion, our research highlights GRg1's potential as an effective adjunct therapy for SALI, primarily by inhibiting apoptosis in alveolar epithelial cells and reducing pro-inflammatory cytokine secretion, thus significantly enhancing the survival rates of CLP mice. These beneficial effects are mediated through targeting AKT1 and activating the PI3K-AKT pathway.

Isolation and anti-neuroinflammation activity of sesquiterpenoids from Artemisia argyi: computational simulation and experimental verification.

BACKGROUND: Artemisia argyi is a traditional herbal medicine belonging to the genus Artemisia that plays an important role in suppressing inflammation. However, the chemical constituents and underlying mechanisms of its therapeutic potential in neuroinflammation are still incompletely understood, and warrant further investigation. METHODS: Several column chromatography were employed to isolate and purify chemical constituents from Artemisia argyi, and modern spectroscopy techniques were used to elucidate their chemical structures. The screening of monomeric compounds with nitric oxide inhibition led to the identification of the most effective bioactive compound, which was subsequently confirmed for its anti-inflammatory capability through qRT‒PCR. Predictions of compound-target interactions were made using the PharmMapper webserver and the TargetNet database, and an integrative protein-protein interaction network was constructed by intersecting the predicted targets with neuroinflammation-related targets. Topological analysis was performed to identify core targets, and molecular docking and molecular dynamics simulations were utilized to validate the findings. The result of the molecular simulations was experimentally validated through drug affinity responsive target stability (DARTS) and Western blot experiments. RESULTS: Seventeen sesquiterpenoids, including fifteen known sesquiterpenoids and two newly discovered guaiane-type sesquiterpenoids (argyinolide S and argyinolide T) were isolated from Artemisia argyi. Bioactivity screening revealed that argyinolide S (AS) possessed the most potent anti-inflammatory activity. However, argyinolide T (AT) showed weak anti-inflammatory activity, so AS was the target compound for further study. AS may regulate neuroinflammation through its modulation of eleven core targets: protein kinase B 1 (AKT1), epidermal growth factor receptor (EGFR), proto-oncogene tyrosine-protein Kinase (FYN), Janus Kinase (JAK) 1, mitogen-activated protein (MAP) Kinase 1,8 and 14, matrix metalloproteinase 9 (MMP9), ras-related C3 botulinum toxin substrate 1 (RAC1), nuclear factor kappa-B p65 (RELA), and retinoid X receptor alpha (RXRA). Molecular dynamics simulations and DARTS experiments confirmed the stable binding of AS to JAK1, and Western blot experiments demonstrated the ability of AS to inhibit the phosphorylation of downstream Signal transducer and activator of transcription 3 (STAT3) mediated by JAK1. CONCLUSIONS: The sesquiterpenoid compounds isolated from Artemisia argyi, exhibit significant inhibitory effects on inflammation in C57BL/6 murine microglia cells (BV-2). Among these compounds, AS, a newly discovered guaiane-type sesquiterpenoid in Artemisia argyi, has been demonstrated to effectively inhibit the occurrence of neuroinflammation by targeting JAK1.

Atomistic Molecular Dynamics Simulations of ABA-Type Polymer Peptide Conjugates: Insights into Supramolecular Structures and their Circular Dichroism Spectra.

A combination of atomistic molecular dynamics (aMD) simulations and circular dichroism (CD) analysis is used to explore supramolecular structures of amphiphilic ABA-type triblock polymer peptide conjugates (PPC). Using the example of a recently introduced PPC with pH- and temperature responsive self-assembling behavior [Otter et al., Macromolecular Rapid Communications 2018, 39, 1800459], this study shows how molecular dynamics simulations of simplified fragment molecules can add crucial information to CD data, which helps to correctly identify the self-assembled structures and monitor the folding/unfolding pathways of the molecules. The findings offer insights into the nature of structural transitions induced by external stimuli, thus contributing to the understanding of the connection of microscopic structures with macroscopic properties.

Apigenin inhibits proliferation and differentiation of cardiac fibroblasts through AKT/GSK3ß signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia miltiorrhiza Bunge (S. miltiorrhiza) is an important Traditional Chinese herbal Medicine (TCM) used to treat cardio-cerebrovascular diseases. Based on the pharmacodynamic substance of S. miltiorrhiza, the aim of present study was to investigate the underlying mechanism of S. miltiorrhiza against cardiac fibrosis (CF) through a systematic network pharmacology approach, molecular docking and dynamics simulation as well as experimental investigation in vitro. MATERIALS AND METHODS: A systematic pharmacological analysis was conducted using the Traditional Chinese Medicine Pharmacology (TCMSP) database to screen the effective chemical components of S. miltiorrhiza, then the corresponding potential target genes of the compounds were obtained by the Swiss Target Prediction and TCMSP databases. Meanwhile, GeneCards, DisGeNET, OMIM, and TTD disease databases were used to screen CF targets, and a protein-protein interaction (PPI) network of drug-disease targets was constructed on S. miltiorrhiza/CF targets by Search Tool for the Retrieval of Interacting Genes/Proteins (STING) database. After that, the component-disease-target network was constructed by software Cytoscape 3.7. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were performed for the intersection targets between drug and disease. The relationship between active ingredient of S. miltiorrhiza and disease targets of CF was assessed via molecular docking and molecular dynamics simulation. Subsequently, the underlying mechanism of the hub compound on CF was experimentally investigated in vitro. RESULTS: 206 corresponding targets to effective chemical components from S. miltiorrhiza were determined, and among them, there were 82 targets that overlapped with targets of CF. Further, through PPI analysis, AKT1 and GSK3ß were the hub targets, and which were both enriched in the PI3K/AKT signaling pathway, it was the sub-pathways of the lipid and atherosclerosis pathway. Subsequently, compound-disease-genes-pathways diagram is constructed, apigenin (APi) was a top ingredients and AKT1 (51) and GSK3ß (22) were the hub genes according to the degree value. The results of molecular docking and dynamics simulation showed that APi has strong affinities with AKT and GSK3ß. The results of cell experiments showed that APi inhibited cells viability, proliferation, proteins expression of α-SMA and collagen I/III, phosphorylation of AKT1 and GSK3ß in MCFs induced by TGFß1. CONCLUSION: Through a systematic network pharmacology approach, molecular docking and dynamics simulation, and confirmed by in vitro cell experiments, these results indicated that APi interacts with AKT and GSK3ß to disrupt the phosphorylation of AKT and GSK3ß, thereby inhibiting the proliferation and differentiation of MCFs induced by TGFß1, which providing new insights into the pharmacological mechanism of S. miltiorrhiza in the treatment of CF.

Simultaneously Enhanced Catalytic Activity and Thermostability of a Baeyer-Villiger Monooxygenase from Oceanicola granulosus by Reshaping the Binding Pocket.

Enzymatic oxygenation of various cyclic ketones into lactones via Baeyer-Villiger monooxygenases (BVMOs) could provide a promising route for synthesizing fragrances and pharmaceutical ingredients. However, unsatisfactory catalytic activity and thermostability restricted their applications in the pharmaceutical and food industries. In this study, we successfully improved the catalytic activity and thermostability of a Baeyer-Villiger monooxygenase (OgBVMO) from Oceanicola granulosus by reshaping the binding pocket. As a result, mutant OgBVMO-Re displayed a 1.0- to 6.4-fold increase in the activity toward branched cyclic ketones tested, accompanied by a 3 °C higher melting point, and a 2-fold longer half-life time (t1/2 (45 °C)). Molecular dynamics simulations revealed that reshaping the binding pocket achieved strengthened motion correlation between amino acid residues, appropriate size of the substrate-binding pocket, beneficial surface characteristics, lower energy barriers, and shorter nucleophilic distance. This study well demonstrated the trade-off between the enzyme activity and thermostability by reshaping the substrate-binding pocket, paving the way for further engineering other enzymes.

One-pot complexation of phytic acid and polyethyleneimine on cellulosic microfibers towards insulative and flame-resistant foam.

Flame resistance is required for the deployment of bio-based materials, especially those forming cellular structures that endow thermal insulation. This study proposes a one-pot strategy to prepare cellular lignocellulosic composites with excellent flame resistance. Lignocellulosic microfibers were used as the substrate onto which a flame-retardant complex consisting of P-containing phytic acid (PA) and N-containing polyethyleneimine (PEI) was formed. Following the prediction of ab initio molecular dynamics simulation, PA and PEI are integrated onto MF-CTMP following a single-step complexation assembly triggered by pH effects. The PA-PEI modified MF-CTMP can be readily transformed into a composite solid foam by dewatering a wet foam followed by oven drying. At the expense of a slightly reduced thermal insulation (thermal conductivity increase from 33.6 ±â€¯0.6 to 40.0 ±â€¯0.6 mW/(m·K)) the presence of PA-PEI complexes significantly improved the mechanical performance of the foam and uniquely endows it with flame resistance. Compared to unmodified MF-CTMP foams, the composite foams showed significant improvement in the Young's, specific compression, and flexural moduli (increased by 13.5, 5.5, and 7.3 folds, respectively), a high oxygen index (up to 40.8 %) and self-extinguishing effects. The results suggest the suitability of the introduced lignocellulosic foam as an alternative to traditional synthetic polymer-based counterparts as well as inorganic matter for insulation, particularly relevant to the building sector.

Proposed dual membrane contact with full-length Osh4.

Membrane contacts sites (MCSs) play important roles in lipid trafficking across cellular compartments and maintain the widespread structural diversity of organelles. We have utilized microsecond long all-atom (AA) molecular dynamics (MD) simulations and enhanced sampling techniques to unravel the MCS structure targeting by yeast oxysterol binding protein (Osh4) in an environment that mimics the interface of membranes with an increased proportion of anionic lipids using CHARMM36m forcefield with additional CUFIX parameters for lipid-protein electrostatic interactions. In a dual-membrane environment, unbiased MD simulations show that Osh4 briefly interacts with both membranes, before aligning itself with a single membrane, adopting a ß-crease-bound conformation similar to observations in a single-membrane scenario. Targeted molecular dynamics simulations followed by microsecond-long AA MD simulations have revealed a distinctive dual-membrane bound state of Osh4 at MCS, wherein the protein interacts with the lower membrane via the ß-crease surface, featuring its PHE-239 residue positioned below the phosphate plane of membrane, while concurrently establishing contact with the opposite membrane through the extended α6-α7 region. Osh4 maintains these dual membrane contacts simultaneously over the course of microsecond-long MD simulations. Moreover, binding energy calculations highlighted the essential roles played by the phenylalanine loop and the α6 helix in dynamically stabilizing dual-membrane bound state of Osh4 at MCS. Our computational findings were corroborated through frequency of contact analysis, showcasing excellent agreement with past experimental cross-linking data. Our computational study reveals a dual-membrane bound conformation of Osh4, providing insights into protein-membrane interactions at membrane contact sites and their relevance to lipid transfer processes.

Electric Field-Enhanced Ion Rejection Rate in Freeze Desalination.

Freeze desalination is an appealing method for seawater desalination through freezing seawater. The percentage of ions in the liquid phase, which is termed ion rejection rate, is a critical factor affecting the performance of freeze desalination. Improving the ion rejection rate is an important topic for freeze desalination. In this work, we investigate the effects of electric fields on the ion rejection rate during the freezing of seawater through molecular dynamics simulations.  It is found that the ion rejection rate increases with increasing electric field strength.  The enhanced ion rejection rate is due to the reduction of the energy barrier at the ice-water interface caused by the electric field, which affects the orientation of water molecules and ion-water interactions. However, the electric field hinders the ice growth rate, which affects the productivity of freeze desalination. Nevertheless, the finding in this work offers a new idea to improve the ion rejection rate. Practically, a trade-off needs to be found to optimize the overall performance of freeze desalination.

Simulation and experimental study of gas-phase diffusion coefficient of selenium dioxide.

Improving the removal effect of selenium in wet flue gas desulfurization system is a key way to reduce the emission of selenium pollutants from coal-fired power plants. In order to clarify the removal mechanism of selenium pollutants in the desulfurization tower, it is necessary to obtain accurate selenium gas-phase diffusion coefficient. In this paper, molecular dynamics simulations were used to carry out theoretical calculations of gas-phase diffusion coefficients of SeO2 (the main form of selenium in coal combustion flue gas). The gas-phase diffusion coefficients of SeO2 in the range of 393 K-433 K were measured by a self-developed heavy metal gas diffusion coefficient testing device to verify the accuracy of the molecular dynamics calculations. Furthermore, the calculated gas-phase diffusion coefficients of SeO2 under typical binary and ternary components were obtained by correcting on the basis of Fuller's formula. Finally, a single-droplet absorption model for SeO2 was constructed and experiments were carried out to compare the effect of the gas-phase diffusion coefficient on the accuracy of the model calculations. The error of the model calculations was reduced from 8.09 % to 1.96 % after the correction. In this study, the gas-phase diffusion coefficient of SeO2 in the low-temperature range of coal-fired flue gas was obtained. This study can provide basic data for the development of selenium migration mechanism and control technology.

A lever hypothesis for Synaptotagmin-1 action in neurotransmitter release.

Neurotransmitter release is triggered in microseconds by Ca2+-binding to the Synaptotagmin-1 C2 domains and by SNARE complexes that form four-helix bundles between synaptic vesicles and plasma membranes, but the coupling mechanism between Ca2+-sensing and membrane fusion is unknown. Release requires extension of SNARE helices into juxtamembrane linkers that precede transmembrane regions (linker zippering) and binding of the Synaptotagmin-1 C2B domain to SNARE complexes through a 'primary interface' comprising two regions (I and II). The Synaptotagmin-1 Ca2+-binding loops were believed to accelerate membrane fusion by inducing membrane curvature, perturbing lipid bilayers or helping bridge the membranes, but SNARE complex binding orients the Ca2+-binding loops away from the fusion site, hindering these putative activities. Molecular dynamics simulations now suggest that Synaptotagmin-1 C2 domains near the site of fusion hinder SNARE action, providing an explanation for this paradox and arguing against previous models of Sytnaptotagmin-1 action. NMR experiments reveal that binding of C2B domain arginines to SNARE acidic residues at region II remains after disruption of region I. These results and fluorescence resonance energy transfer assays, together with previous data, suggest that Ca2+ causes reorientation of the C2B domain on the membrane and dissociation from the SNAREs at region I but not region II. Based on these results and molecular modeling, we propose that Synaptotagmin-1 acts as a lever that pulls the SNARE complex when Ca2+ causes reorientation of the C2B domain, facilitating linker zippering and fast membrane fusion. This hypothesis is supported by the electrophysiological data described in the accompanying paper.

Repurposing of Drug Bank Compounds against Plasmodium falciparum Dihydroorotate Dehydrogenase as novel anti malarial drug candidates by Computational approaches.

This study aimed to repurpose Drug Bank Compounds against P. falciparum Dihydroorotate dehydrogenase (Pf-DHODH)a potential molecular target for antimalarial drug development due to its vital role in P. falciparum survival. Initially, the MATGEN server was used to screen drugs against Pf-DHODH (PDB ID 6GJG), followed by revalidating the results through docking by Autodock Vina through PyRx. Based on the docking results, three drugs namely, Talnifumate, Sulfaphenazole, and (3S)-N-[(2S)-1-[2-(1H-indol-3-yl)ethylamino]-1-oxopropan-2-yl]-1-(4-methoxyphenyl)-5-oxopyrrolidine-3-carboxamide-were subjected to molecular dynamics simulation for 100 ns. Molecular dynamics simulation results indicate that (3S)-N-[(2S)-1-[2-(1H-indol-3-yl)ethylamino]-1-oxopropan-2-yl]-1-(4-methoxyphenyl)-5-oxopyrrolidine-3-carboxamide- and Sulfaphenazole may target Pf-DHODH by forming a stable protein-ligand complex as they showed better free binding energy -130.58 kJ/mol, and -79.84 kJ/mol, respectively as compared to the free binding energy 116.255 kJ/mol of the reference compound; 3,6-dimethyl- ~ {N}-[4-(trifluoromethyl)phenyl]-[1,2]oxazolo[5,4-d]pyrimidin-4-amine. Although the studied compounds are drugs, still we applied Lipinski's rules and ADMET analysis that reconfirmed that these drugs have favorable drug-like properties. In conclusion, the results of the study show that Talniflumate and Sulfaphenazole may be potential antimalarial drug candidates.The derivatives of these drugs could be designed and tested to develop better drugs against Plasmodium species.

Triple Effects of the Physicochemical Interaction between Water and Copper and Their Influence on Microcutting.

Water has been recognized in promoting material removal, traditionally ascribed to friction reduction and thermal dissipation. However, the physicochemical interactions between water and the workpiece have often been overlooked. This work sheds light on how the physicochemical interactions that occur between water (H2O) and copper (Cu) workpiece influence material deformations during the cutting process. ReaxFF molecular dynamics simulations were employed as the primary method to study the atomistic physical and chemical interactions between the applied medium and the workpiece. Upon contact with the Cu surface, H2O dissociated into OH- ions, H+ ions, and traces of O2- ions. The OH- and O2- ions chemically reacted with Cu to form bonds that weakened the Cu-Cu bonds by elongation, while the H+ ions gained electrons and diffused into the Cu lattice as H- ions. The weakening of surface Cu bonds promoted plastic deformation and reduced the difficulty of material removal. Meanwhile, further addition of H2O molecules saw a plateau in hydrolysis and more dominance of H2O physical adsorption on Cu, which weakens the elongation of Cu-Cu bonds. While the ideal case for atomic-scale material removal was found with an optimal number of 240 H2O molecules, the presented Cu material state with more H2O molecules could account for the observations in microcutting. The constricted nature of physical adsorption and hydrogen ion diffusion in the surface layer prevented the propagation of dislocations through the surface, which subsequently caused pinning points to be closer together during chip formation as observed by smaller chip fold widths on the microscale. Theoretical and experimental analysis identified the importance of accounting for physicochemical interactions between surface media and the workpiece when considering material deformations at micronanoscale.

Investigating the applicability of novel hydrate dissociation inhibitors in oilwell cement through molecular simulations.

In the field of hydrate formation cementing, the method of developing the low hydration exothermic cement systems cannot effectively solve the problem of hydrate dissociation caused by the hydration heat release of cement. Therefore, we proposed a new approach to address this issue by employing cement additives that can effectively delay the dissociation of hydrate. In our previous work, we designed a novel hydrate dissociation inhibitor, PVCap/dmapma, however, its applicability with cement slurry remains unverified. In this study, we established a more realistic model of oilwell cement gel based on experimental data. Additionally, we investigated the potential effects of PVCap/dmapma on the microstructure and mechanical properties of cement gel through molecular simulations. The results suggest that PVCap/dmapma has no negative effect on the performance of cement slurry compared to Lecithin. By adding PVCap/dmapma to cement slurry, the problem of cementing in hydrate formations is expected to be solved.

In silico discovery of potential PPI inhibitors for anti-lung cancer activity by targeting the CCND1-CDK4 complex via the P21 inhibition mechanism.

Non-Small Cell Lung Cancer (NSCLC) is a prevalent and deadly form of lung cancer worldwide with a low 5-year survival rate. Current treatments have limitations, particularly for advanced-stage patients. P21, a protein that inhibits the CCND1-CDK4 complex, plays a crucial role in cell proliferation. Computer-Aided Drug Design (CADD) based on pharmacophores can screen and design PPI inhibitors targeting the CCND1-CDK4 complex. By analyzing known inhibitors, key pharmacophores are identified, and computational methods are used to screen potential PPI inhibitors. Molecular docking, pharmacophore matching, and structure-activity relationship studies optimize the inhibitors. This approach accelerates the discovery of CCND1-CDK4 PPI inhibitors for NSCLC treatment. Molecular dynamics simulations of CCND1-CDK4-P21 and CCND1-CDK4 complexes showed stable behavior, comprehensive sampling, and P21's impact on complex stability and hydrogen bond formation. A pharmacophore model facilitated virtual screening, identifying compounds with favorable binding affinities. Further simulations confirmed the stability and interactions of selected compounds, including 513457. This study demonstrates the potential of CADD in optimizing PPI inhibitors targeting the CCND1-CDK4 complex for NSCLC treatment. Extended simulations and experimental validations are necessary to assess their efficacy and safety.

Mode of molecular interaction of triterpenoid saponin ginsenoside Rh2 with membrane lipids in liquid-disordered phases.

Ginsenoside Rh2 (Rh2) is a ginseng saponin comprising a triterpene core and one unit of glucose and has attracted much attention due to its diverse biological activities. In the present study, we used small-angle X-ray diffraction, solid-state NMR, fluorescence microscopy, and MD simulations to investigate the molecular interaction of Rh2 with membrane lipids in the liquid-disordered (Ld) phase mainly composed of palmitoyloleoylphosphatidylcholine compared with those in liquid-ordered (Lo) phase mainly composed of sphingomyelin and cholesterol. The electron density profiles determined by X-ray diffraction patterns indicated that Rh2 tends to be present in the shallow interior of the bilayer in the Ld phase, while Rh2 accumulation was significantly smaller in the Lo phase. Order parameters at intermediate depths in the bilayer leaflet obtained from 2H NMR spectra and MD simulations indicated that Rh2 reduces the order of the acyl chains of lipids in the Ld phase. The dihydroxy group and glucose moiety at both ends of the hydrophobic triterpene core of Rh2 cause tilting of the molecular axis relative to the membrane normal, which may enhance membrane permeability by loosening the packing of lipid acyl chains. These features of Rh2 are distinct from steroidal saponins such as digitonin and dioscin, which exert strong membrane-disrupting activity.

Catalpol reduced LPS induced BV2 immunoreactivity through NF-κB/NLRP3 pathways: an in Vitro and in silico study.

Background: Ischemic Stroke (IS) stands as one of the primary cerebrovascular diseases profoundly linked with inflammation. In the context of neuroinflammation, an excessive activation of microglia has been observed. Consequently, regulating microglial activation emerges as a vital target for neuroinflammation treatment. Catalpol (CAT), a natural compound known for its anti-inflammatory properties, holds promise in this regard. However, its potential to modulate neuroinflammatory responses in the brain, especially on microglial cells, requires comprehensive exploration. Methods: In our study, we investigated into the potential anti-inflammatory effects of catalpol using lipopolysaccharide (LPS)-stimulated BV2 microglial cells as an experimental model. The production of nitric oxide (NO) by LPS-activated BV2 cells was quantified using the Griess reaction. Immunofluorescence was employed to measure glial cell activation markers. RT-qPCR was utilized to assess mRNA levels of various inflammatory markers. Western blot analysis examined protein expression in LPS-activated BV2 cells. NF-κB nuclear localization was detected by immunofluorescent staining. Additionally, molecular docking and molecular dynamics simulations (MDs) were conducted to explore the binding affinity of catalpol with key targets. Results: Catalpol effectively suppressed the production of nitric oxide (NO) induced by LPS and reduced the expression of microglial cell activation markers, including Iba-1. Furthermore, we observed that catalpol downregulated the mRNA expression of proinflammatory cytokines such as IL-6, TNF-α, and IL-1ß, as well as key molecules involved in the NLRP3 inflammasome and NF-κB pathway, including NLRP3, NF-κB, caspase-1, and ASC. Our mechanistic investigations shed light on how catalpol operates against neuroinflammation. It was evident that catalpol significantly inhibited the phosphorylation of NF-κB and NLRP3 inflammasome activation, both of which serve as upstream regulators of the inflammatory cascade. Molecular docking and MDs showed strong binding interactions between catalpol and key targets such as NF-κB, NLRP3, and IL-1ß. Conclusion: Our findings support the idea that catalpol holds the potential to alleviate neuroinflammation, and it is achieved by inhibiting the activation of NLRP3 inflammasome and NF-κB, ultimately leading to the downregulation of pro-inflammatory cytokines. Catalpol emerges as a promising candidate for the treatment of neuroinflammatory conditions.

Network Pharmacology and Molecular Docking Integrated with Molecular Dynamics Simulations Investigate the Pharmacological Mechanism of Yinchenhao Decoction in the Treatment of Non-alcoholic Fatty Liver Disease.

BACKGROUND: Non-Alcoholic Fatty Liver Disease (NAFLD) has become a significant health and economic burden globally. Yinchenhao decoction (YCHD) is a traditional Chinese medicine formula that has been validated to exert therapeutic effects on NAFLD. OBJECT: The current study aimed to explore the pharmacological mechanisms of YCHD on NAFLD and further identify the potential active compounds acting on the main targets. METHODS: Compounds in YCHD were screened and collected from TCMSP and published studies, and their corresponding targets were obtained from the SWISS and SEA databases. NAFLD-related targets were searched in the GeneCards and DisGeNet databases. The "compound- intersection target" network was constructed to recognize the key compounds. Moreover, a PPI network was constructed to identify potential targets. GO and KEGG analyses were performed to enrich the functional information of the intersection targets. Then, molecular docking was used to identify the most promising compounds and targets. Finally, molecular dynamics (MD) simulations were performed to verify the binding affinity of the most potential compounds with the key targets. RESULTS: A total of 53 compounds and 556 corresponding drug targets were collected. Moreover, 2684 NAFLD-related targets were obtained, and 201 intersection targets were identified. Biological processes, including the apoptotic process, inflammatory response, xenobiotic metabolic process, and regulation of MAP kinase activity, were closely related to the treatment of NAFLD. Metabolic pathways, non-alcoholic fatty liver disease, the MAPK signaling pathway, and the PI3K-Akt signaling pathway were found to be the key pathways. Molecular docking showed that quercetin and isorhamnetin were the potential active compounds, while AKT1, IL1B, and PPARG were the most promising targets. MD simulations further verified that the binding of PPARG-isorhamnetin (-35.96 ± 1.64 kcal/mol) and AKT1-quercetin (-31.47 ± 1.49 kcal/mol) was due to their lowest binding free energy. CONCLUSION: This study demonstrated that YCHD exerts therapeutic effects for the treatment of NAFLD through multiple targets and pathways, providing a theoretical basis for further pharmacological research on the potential mechanisms of YCHD in NAFLD.

Mechanisms underlying the therapeutic effects of Gang Huo Qing wen granules in the treatment of influenza based on network pharmacology, molecular docking and molecular dynamics.

Influenza (Flu) is a severe health, medical, and economic problem, but no medication that has excellent outcomes and lowers the occurrence of these problems is now available. GanghuoQingwenGranules (GHQWG) is a common Chinese herbal formula for the treatment of influenza (flu). However, its methods of action remain unknown. We used network pharmacology, molecular docking, and molecular dynamics simulation techniques to investigate the pharmacological mechanism of GHQWG in flu. TCMSP and various types of literature were used to obtain active molecules and targets of GHQWG. Flu-related targets were found in the Online Mendelian Inheritance in Man (OMIM) database, the DisFeNET database, the Therapeutic Target Database (TTD), and the DrugBank database. To screen the key targets, a protein-protein interaction (PPI) network was constructed. DAVID was used to analyze GO and KEGG pathway enrichment. Target tissue and organ distribution was assessed. Molecular docking was used to evaluate interactions between possible targets and active molecules. For the ideal core protein-compound complexes obtained using molecular docking, a molecular dynamics simulation was performed. In total, 90 active molecules and 312 GHQWG targets were discovered. The PPI network's topology highlighted six key targets. GHQWG's effects are mediated via genes involved in inflammation, apoptosis, and oxidative stress, as well as the TNF and IL-17 signaling pathways, according to GO and KEGG pathway enrichment analysis. Molecular docking and molecular dynamics simulations demonstrated that the active compounds and tested targets had strong binding capabilities. This analysis accurately predicts the effective components, possible targets, and pathways involved in GHQWG flu treatment. We proposed a novel study strategy for future studies on the molecular processes of GHQWG in flu treatment. Furthermore, the possible active components provide a dependable source for flu drug screening.

Mechanistic insights into the conversion of flavin adenine dinucleotide (FAD) to 8-formyl FAD in formate oxidase: a combined experimental and in-silico study.

Formate oxidase (FOx), which contains 8-formyl flavin adenine dinucleotide (FAD), exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-choline (GMC) oxidoreductase enzymes that contain only the standard FAD cofactor. The FOx-mediated conversion of FAD to 8-formyl FAD results in an approximate 10-fold increase in formate oxidase activity. However, the mechanistic details underlying the autocatalytic formation of 8-formyl FAD are still not well understood, which impedes further utilization of FOx. In this study, we employ molecular dynamics simulation, QM/MM umbrella sampling simulation, enzyme activity assay, site-directed mutagenesis, and spectroscopic analysis to elucidate the oxidation mechanism of FAD to 8-formyl FAD. Our results reveal that a catalytic water molecule, rather than any catalytic amino acids, serves as a general base to deprotonate the C8 methyl group on FAD, thus facilitating the formation of a quinone-methide tautomer intermediate. An oxygen molecule subsequently oxidizes this intermediate, resulting in a C8 methyl hydroperoxide anion that is protonated and dissociated to form OHC-RP and OH-. During the oxidation of FAD to 8-formyl FAD, the energy barrier for the rate-limiting step is calculated to be 22.8 kcal/mol, which corresponds to the required 14-hour transformation time observed experimentally. Further, the elucidated oxidation mechanism reveals that the autocatalytic formation of 8-formyl FAD depends on the proximal arginine and serine residues, R87 and S94, respectively. Enzymatic activity assay validates that the mutation of R87 to lysine reduces the kcat value to 75% of the wild-type, while the mutation to histidine results in a complete loss of activity. Similarly, the mutant S94I also leads to the deactivation of enzyme. This dependency arises because the nucleophilic OH- group and the quinone-methide tautomer intermediate are stabilized through the noncovalent interaction provided by R87 and S94. These findings not only explain the mechanistic details of each reaction step but also clarify the functional role of R87 and S94 during the oxidative maturation of 8-formyl FAD, thereby providing crucial theoretical support for the development of novel flavoenzymes with enhanced redox properties.

Immunoinformatics and structural aided approach to develop multi-epitope based subunit vaccine against Mycobacterium tuberculosis.

Tuberculosis is a highly contagious disease caused by Mycobacterium tuberculosis (Mtb), which is one of the prominent reasons for the death of millions worldwide. The bacterium has a substantially higher mortality rate than other bacterial diseases, and the rapid rise of drug-resistant strains only makes the situation more concerning. Currently, the only licensed vaccine BCG (Bacillus Calmette-Guérin) is ineffective in preventing adult pulmonary tuberculosis prophylaxis and latent tuberculosis re-activation. Therefore, there is a pressing need to find novel and safe vaccines that provide robust immune defense and have various applications. Vaccines that combine epitopes from multiple candidate proteins have been shown to boost immunity against Mtb infection. This study applies an immunoinformatic strategy to generate an adequate multi-epitope immunization against Mtb employing five antigenic proteins. Potential B-cell, cytotoxic T lymphocyte, and helper T lymphocyte epitopes were speculated from the intended proteins and coupled with 50 s ribosomal L7/L12 adjuvant, and the vaccine was constructed. The vaccine's physicochemical profile demonstrates antigenic, soluble, and non-allergic. In the meantime, docking, molecular dynamics simulations, and essential dynamics analysis revealed that the multi-epitope vaccine structure interacted strongly with Toll-like receptors (TLR2 and TLR3). MM-PBSA analysis was performed to ascertain the system's intermolecular binding free energies accurately. The immune simulation was applied to the vaccine to forecast its immunogenic profile. Finally, in silico cloning was used to validate the vaccine's efficacy. The immunoinformatics analysis suggests the multi-epitope vaccine could induce specific immune responses, making it a potential candidate against Mtb. However, validation through the in-vivo study of the developed vaccine is essential to assess its efficacy and immunogenicity profile, which will assure active protection against Mtb.