MD Simulations Research Articles

Drug Binding to Partially Unfolded Serum Albumin: Insights into Nonsteroidal Anti-Inflammatory Drug Naproxen-BSA Interactions from Spectroscopic and MD Simulation Studies.

Understanding the binding details of a small-molecule drug to a protein in its partially unfolded state is important for drug delivery as it provides insight into the overall drug-binding ability of the protein, even when the majority of binding pockets in its unfolded state are impaired. The interaction of partially unfolded proteins with drugs remains poorly understood due to a lack of structural information on proteins in their partially unfolded states. Here, we studied the interaction between serum albumin (bovine serum albumin (BSA) as a model system), an abundant protein in blood serum that is an effective carrier for numerous known drugs, and a nonsteroidal anti-inflammatory drug (NSAID) naproxen (NPS) using various spectroscopic and computational methods. Molecular dynamics simulations starting from the drug-unbound state and performed at physiological and higher temperatures revealed novel hydrophobic sites on the BSA surface. We analyzed the BSA-NPS interaction in the presence and absence of the cationic organized assembly CTAB and two oligosaccharides (β-CD and 2-HP-β-CD) at different excitation wavelengths. The solvation dynamics of BSA under NPS-bound conditions became ∼4.6% faster. Oligosaccharides were found to increase the solubility of NPS by providing a hydrophobic environment for the formation of inclusion complexes through host-guest interactions. These findings provide a comprehensive overview and uncover the binding model and mechanism of interaction of NPS with BSA, revealing hydrophobic and electrostatic interactions and hydrogen bonds required for BSA to bind NPS at these noncanonical sites. The molecular-level understanding of the binding mechanism of commonly used NSAIDs like NPS with partially unfolded BSA will be useful in designing pharmaceutically important molecules with efficient loading and delivery properties.

Modelling DNA-ligand interactions through variable Force Field based MD Simulations

Modelling DNA-ligand interactions through variable Force Field based MD Simulations

Understanding the Functional Dynamics of the TokK Enzyme in Carbapenem Biosynthesis via MD Simulations and QM/MM Calculations.

Due to the recent surge in antibiotic resistance, developing novel antibiotics is the demand of the time, and thus, a precise understanding of the catalytic mechanisms of enzymes involved in antibiotic biosynthesis becomes crucial. Here, we present a comprehensive investigation into the catalytic mechanism of TokK, a freshly characterized B12-dependent RSMT enzyme that plays an important role in carbapenem biosynthesis. Using MD simulations, we show how the plasticity of the active site facilitates substrate recognition while the quantum mechanics/molecular mechanics calculations provide a detailed mechanistic understanding of the methyl transfer process, elucidating stereochemical preferences. Notably, we demonstrate the indispensable role of Trp215 in orchestrating the proper orientation of the 5'-dA• radical for efficient substrate methylation, which strongly correlates with the previous findings where the mutation of Trp215 has severely affected the enzyme activity.

Synthesis, Characterization and Assessment of Antioxidant and Melanogenic Inhibitory Properties of Edaravone Derivatives.

A series of edaravone derivatives and the corresponding Cu(II) complexes were synthesized and characterized using spectroscopic and analytical techniques such as IR, UV, NMR and elemental analysis. Antioxidant activities of all compounds were examined using free radical scavenging methods such as hydrogen peroxide scavenging activity (HPSA), 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2-2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonate) (ABTS) assays. All of the tested compounds exhibited good antioxidant activity. Further, the frontier orbital energy levels, as well as various chemical properties, were determined using the density functional theory (DFT) calculations. The MEP maps of all of the derivatives were plotted to identify the nucleophilic and electrophilic reactive sites. Further, binding energies of all of the organic compounds with the protein tyrosinase was investigated to determine their potential anti-melanogenic applications. The selected ligand, L6 was subjected to molecular dynamics simulation analysis to determine the stability of the ligand-protein complex. The MD simulation was performed (150 ns) to estimate the stability of the tyrosinase-L6 complex. Other key parameters, such as, RMSD, RMSF, Rg, hydrogen bonds, SASA and MMPBSA were also analyzed to understand the interaction of L6 with the tyrosinase protein.

Discovery of Small Molecule Inhibitors Targeting CTNNB1 (β-catenin) for Endometrial cancer: Employing 3D QSAR, Drug-Likeness Assessment, ADMET Predictions, Molecular Docking and Simulation.

Endometrial carcinoma (EC) is a type of cancer that originates in the lining of the uterus, known as the endometrium. It is associated with various treatment options such as surgery, radiation therapy, chemotherapy, and hormone therapy, each presenting unique challenges and limitations. Beta-catenin, a protein involved in the development and progression of several cancers, including EC, plays a crucial role. Abnormal beta-catenin signaling is often linked to the emergence of specific EC subtypes, affecting tumor growth and invasion. The study's objective is to identify compounds targeting the beta-catenin protein for treating endometrial cancer (EC) using in silico drug design. Our approach includes molecular docking to evaluate binding affinities, ADME profiling for pharmacokinetic properties, toxicity assessments, and molecular dynamics simulations to assess compound stability and interactions. Approximately one thousand anti-cancer phytochemicals were sourced from PubChem and subjected to molecular docking simulations against the beta-catenin protein. The compounds were evaluated based on their binding affinities, with the top five selected for further analysis. These five molecules underwent toxicity and ADME profiling. The Prediction of Activity Spectra for Substances (PASS) tool was used to identify compounds targeting CTNNB1. Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were employed to establish quantitative structure-activity relationship (QSAR) models for the five CTNNB1 antagonist molecules. The selected five compounds, namely Pazopanib, Binimetinib, Telatinib, 4-(2,3-Dihydrobenzo[ b][1,4]dioxin-6-yl)-3-((5-nitrothiazol-2-yl)thio)-1H-1,2,4-triazol-5(4H)-one, and Ribavirin, demonstrated efficacy against CTNN1. MD simulations of the docked complexes confirmed the stability of these drugs in binding to the target protein. All five molecules showed promising safety and effectiveness profiles according to their ADME and toxicity evaluations. Through a comprehensive screening process employing in silico drug design methods, this study successfully identified five potential human anticancer drug candidates targeting the beta-catenin protein. These findings offer a foundation for further experimental validation and development towards the treatment of EC.

Utilizing Cinnamomum verum (a culinary spice), as a functional food ingredient ameliorating hypercholesterolemia: In-vivo, in-vitro, and in-silico multi-model analysis

Hypercholesterolemia involves elevated levels of total cholesterol (T.C.), low-density lipoprotein (LDL), very low-density lipoprotein (VLDL), and reduced high-density lipoprotein (HDL), linked to sedentary lifestyles, irregular eating habits, high-fat diets, type 2 diabetes, and various hereditary and environmental factors. It poses significant health risks, including high blood pressure, atherosclerosis, fatty liver, and cardiovascular disease. This study aimed to evaluate the lipid-lowering effects of ethanolic extract of Cinnamon in hypercholesterolemia-induced rats using in-vivo, in-vitro, and in-silico approaches. In the present work, thirty female Albino rats were divided into six groups: Group I (basal diet), Group II (high-fat diet), Group III (statin treatment), and Groups IV-VI (high-fat diet with different doses of cinnamon extract). Serum lipid profiles were measured after four weeks of treatment. In-vitro HMG-CoA reductase inhibition was assessed. In-silico studies, including molecular docking, ADME analysis, toxicity prediction, and molecular dynamics (M.D.) simulations, were conducted. DFT studies with MESP/HOMO/LUMO analysis provided electronic property insights. High-fat diet increased body weight and serum lipid levels in rats. Cinnamon extract significantly reduces body weight and serum LDL, VLDL, and triglycerides while increasing HDL levels. Histopathology showed reduced fat accumulation and normal liver morphology in cinnamon-treated groups. In-vitro analysis revealed significant HMG-CoA reductase inhibition by cinnamon extract. In-silico docking confirmed strong binding affinities of cinnamon compounds to HMG-CoA reductase. ADME analysis and toxicity prediction indicated favorable pharmacokinetics. MD simulations showed stable ligand-protein complexes, and DFT studies highlighted the electronic properties of active compounds. To conclude, the cinnamon ethanolic extract demonstrated effective lipid-lowering properties in hypercholesterolemic rats, supported by in-vitro and in-silico analyses, suggesting its potential as a therapeutic agent for hypercholesterolemia. Further research is needed to explore underlying mechanisms and clinical outcomes.

Amyloid-like Aggregation Propensities of Metabolites -Homogentisic acid, N-Acetyl aspartic acid and Isovaleric acid.

The transformation of metabolites into amyloidogenic aggregates represent an intriguing dimension in the pathophysiology of metabolic disorders, including alkaptonuria, canavan disease, and isovaleric acidemia. Central to this phenomenon are the metabolites homogentisic acid (HA), N-acetyl aspartic acid (NAA), and isovaleric acid (IVA), which we found, weave an intricate network of self-assembled structures. Leveraging an array of microscopy techniques, we traced the morphological behavior of these assemblies that exhibit concentration and time-dependent morphological transitions from isolated globules to clustered aggregates. MD simulation studies suggest significant role of hydrogen bonding interactions in the aggregation process. While displaying strong amyloidogenic propensity in solution, these aged aggregates were significantly cytotoxic to mouse neural N2a cell lines. In vivo effect in Caenorhabditis elegans (C. elegans) nematode further validated cytotoxicity of aggregates. Our findings provide fresh insights to amyloidogenic nature of HA, NAA, and IVA aggregates and their possible role in associated metabolic disorders such as alkaptonuria, canavan disease and isovaleric acidemia.

Exploring flavonoids as potential inhibitors for Enterococcus faecium nicotinate nucleotide adenylyltransferase: An integration of in silico with empirical studies

Nicotinate nucleotide adenylyltransferase (NNAT) emerges as a promising target for drug development, given its pivotal role in the nicotinate nucleotide biosynthesis pathway crucial for bacterial survival. Using computational modelling and fluorescence methods, this study screened a library of 1427 compounds from MedChemExpress as potential inhibitors against Enterococcus faecium NNAT (EfNNAT). We employed Maestro's Glide HTVS, SP, and XP docking protocols to identify compounds with high binding affinity. The screening utilised two models of the EfNNAT crystal structure: energy-minimised and 100-ns MD-simulated model, hypothesising conformational change that may influence ligand selectivity in the HTVS study. Our findings revealed variations between the minimised versus MD-simulated models, impacting ligand selectivity by each model post-HTVS. However, quercetin 3-O-beta-d-glucose-7-O-beta-d-gentiobioside (QGG) appeared to be the top inhibitor selected by both models after more rigorous docking using Maestro-implemented SP and XP. A 500-ns all-atom MD simulation of the EfNNAT:ligand complex revealed that the compound interacts mainly through hydrogen bonding and water bridges. Validation of the in-silico studies via molecular spectroscopy confirmed QGG's binding to EfNNAT by displacing the fluorescent probes from the protein binding site. Although the binding demonstrated minimal impact on the thermal stability of EfNNAT as deduced from the thermal shift assay, ITC showed that QGG exhibits moderate binding affinity for EfNNAT, with the interaction being spontaneous and thermodynamically favourable. Insights gleaned from this study offer a mechanistic basis that could be leveraged to develop new leads. Expansion of the compound library holds promise for identifying inhibitors with enhanced affinity for EfNNAT.

Abstract B010: Design of personalized neoantigen mRNA vaccines against breast cancer patients in Pakistan based on next-generation sequencing data

Abstract Breast cancer is the most common malignancy among women in Pakistan, with an outrageous increase in the incidence and mortality rate. Because of decreased effectiveness and lower survival rates, there is an urgent need for developing novel, personalized vaccines with enhanced activity. The development of a personalized mRNA vaccine illustrates a promising approach to combating the prevalence of breast cancer. We used the whole exome sequencing (WES) data of Pakistani tumor samples to predict immunogenic neoantigens. The WES data was aligned to the GRCM39 reference genome using BWA. The genome analysis toolkit (GATK) was employed for variant calling to identify somatic mutations. Using immune-bioinformatics tools, we predicted cytotoxic CD8+ T cell epitopes and helper CD4+ T cell epitopes within the identified neoantigens and designed vaccines by assembling the fusion of CTL neoepitopes, helper sequences, and adjuvants together with linkers. The Insilco ImmSim algorithm was used to examine the immune responses of the vaccine. The vaccine design was further explored by checking its simulation effect on the immune system, its physiochemical characteristics, its binding to specific immune system molecules, and cloning into an appropriate vector. In addition, molecular docking, MD simulation, and binding free energies were performed to investigate the interaction mechanism between the construct vaccine and Toll-like receptors (TLR2, TLR4, TLR7, and TLR9).By examining these factors, we aimed to ensure the vaccine's safety, stability, and ability to bind tightly to specific immune cells (class I MHC molecules), ultimately triggering a comprehensive immune response against breast cancer cells. Citation Format: Kayode Yomi Raheem, Muhammad Muddassar. Design of personalized neoantigen mRNA vaccines against breast cancer patients in Pakistan based on next-generation sequencing data [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr B010.

Synthesis, cytotoxic evaluation, and in silico studies of novel benzenesulfonamide-thiazolidinone derivatives against colorectal carcinoma

Thirteen new benzenesulfonamide derivatives containing 4-thiazolidinone were synthesized. Their cytotoxic properties were tested on colorectal carcinoma (HT-29, DLD-1, COLO-205) and normal colon fibroblast (CCD-986Sk) cell lines. Compounds 3l (IC50=114.62 µM), 3a (IC50=25.82 µM), 3k (IC50=30.99 µM), and 3i (IC50=62.94 µM) showed the highest cytotoxicity on HT-29, DLD-1, COLO-205, and CCD-986Sk cell lines, respectively. Compound 3e (IC50=1075.4 µM) exhibited the least cytotoxicity against CCD-986Sk. The apoptosis profile of 5-FU at 5 µM was similar to that of compound 3e at 30 µM. Molecular docking and MD simulations showed stable interactions for compounds 3i and 3e. Molecular docking and MD simulations revealed that compounds 3i and 3e exhibit stable interactions with key cancer-related protein targets, particularly TGFβ2. These interactions suggest their potential as therapeutic agents. The predicted ADME parameters further highlighted compounds 3e and 3i for their favorable lipophilicity, solubility, permeability, and oral absorption profiles, supporting their promise as candidates for future drug development.

Elaboration and investigation of formulated polymer composites as some potential anticorrosion coatings for brass surface in 3.5% NaCl solution: theoretical approaches

ABSTRACT Polymer composite derivatives [(TGEMM/MDA/0%, TGEMM/MDA/6%, TGP/MDA/0%, and TGP/MDA/6%)] were elaborated through using tetraglycidyl ether of methyl-α-D-mannopyranoside (TGEMM) and 2,3,4,5-tetraglycidyloxy pentanal (TGP) as matrices, methylene dianiline (MDA) as curing agent and glass system (1-x) (2Bi2O3 - B2O3) - x BaO (with x = 0% and 6%) as filler. Different composites prepared were investigated as some potential anticorrosive protection coatings for brass electrode in sodium chloride medium (3.5% NaCl). Polymer composite coatings were characterized using FTIR spectroscopy, SEM/EDS, EFM, PDP, and EIS techniques. The anticorrosive protection efficiencies increase with increasing charge incorporated in composites, in the following order: TGP/MDA/0% < TGEMM/MDA/0% < TGP/MDA/6% < TGEMM/MDA/6% and the later achieving a maximum of 90.67% (EFM), 95.45% (PDP), and 97.86% (EIS), respectively. SEM/EDS characterization suggests a smooth surface in the presence of composite derivatives, which is more significant in the presence of TGEMM/MDA/6%. DFT calculations and MD simulations confirm the data obtained by experimental study and offer insights into its structural properties.

Comparative study of stability and activity of wild-type and mutant human carbonic anhydrase II enzymes using molecular dynamics and docking simulations

The human carbonic anhydrase II (HCA II) enzyme is a cytosolic protein that catalyzes the reversible hydration of carbon dioxide (CO2). Considering the critical role of the HCA II and the effects of some mutations on the activity and stability of the enzyme in humans, several computational methods are used to study the structure and dynamics of the wild-type and the mutant enzymes with three ligands, CO2, 4-nitrophenyl acetate and acetazolamide. Our results of MD simulation of a wild-type enzyme with 4-nitrophenyl acetate show that it created essential effects on the fluctuation of this enzyme and made it more unstable and less compact than the same enzyme without ligand. In the MD of the mutant enzyme with 4-nitrophenyl acetate ligand, no significant difference is observed between with and without ligand. The affinity of the wild-type enzyme to the 4-nitrophenyl acetate is notably higher than the mutant enzyme with the same ligand. Furthermore, results showed that wild-type and mutant enzymes with CO2 are more favorable in stability and flexibility than the same enzymes without ligand. The MD results of wild-type with acetazolamide indicate instability compare without ligand, but in MD of mutant enzyme with acetazolamide show that it more stable and compact than the same enzyme without ligand. Finally, Comparing protein trajectories to assess the impact of ligands on the stability and activity of HCA II enzymes can have medical applications and can in the engineering and design of new variants of carbonic anhydrase enzyme.

Lipophilic compounds restore function to neurodevelopmental-associated KCNQ3 mutations

A major driver of neuronal hyperexcitability is dysfunction of K+ channels, including voltage-gated KCNQ2/3 channels. Their hyperpolarized midpoint of activation and slow activation and deactivation kinetics produce a current that regulates membrane potential and impedes repetitive firing. Inherited mutations in KCNQ2 and KCNQ3 are linked to a wide spectrum of neurodevelopmental disorders (NDDs), ranging from benign familial neonatal seizures to severe epileptic encephalopathies and autism spectrum disorders. However, the impact of these variants on the molecular mechanisms underlying KCNQ3 channel function remains poorly understood and existing treatments have significant side effects. Here, we use voltage clamp fluorometry, molecular dynamic simulations, and electrophysiology to investigate NDD-associated variants in KCNQ3 channels. We identified two distinctive mechanisms by which loss– and gain–of function NDD-associated mutations in KCNQ3 affect channel gating: one directly affects S4 movement while the other changes S4-to-pore coupling. MD simulations and electrophysiology revealed that polyunsaturated fatty acids (PUFAs) primarily target the voltage-sensing domain in its activated conformation and form a weaker interaction with the channel’s pore. Consistently, two such compounds yielded partial and complete functional restoration in R227Q- and R236C-containing channels, respectively. Our results reveal the potential of PUFAs to be developed into therapies for diverse KCNQ3-based channelopathies.

Negative Charges, Not Necessary Phosphorylation, are Required for Ligand Recognition by 14-3-3 Proteins.

Protein-protein interactions involving 14-3-3 proteins regulate various cellular activities in normal and pathological conditions. These interactions have mostly been reported to be phosphorylation-dependent, but the 14-3-3 proteins also interact with unphosphorylated proteins. In this work, we investigated whether phosphorylation is required, or, alternatively, whether negative charges are sufficient for 14-3-3ε binding. We substituted the pThr residue of pT(502-510) peptide by residues with varying number of negative charges, and investigated binding of the peptides to 14-3-3ε using MD simulations and biophysical methods. We demonstrated that at least one negative charge is required for the peptides to bind 14-3-3ε while phosphorylation is not necessary, and that two negative charges are preferable for high affinity binding.

Adenosyl derivatives as a potential inhibitors of NS3 protease of Japanese encephalitis virus (JEV): In silico molecular insight into therapeutic discovery

The genus Flavivirus NS3 a non-structural protein of Japanes Encephalitis Virus (JEV), a serious deadly human pathogen responsible for several deaths in South East Asia, consists of helicase/NTPase domain forming a cleft known for their role in the enzymatic activity and viral replication represented biological relevance. S-Adenosyl derivatives act as powerful inhibitors in viral replication in NS5 of the same genus as of NS3, provide a powerful base to test its effectiveness in NS3 protein due to the compatibility of both proteins. The MM-GBSA (Molecular Mechanics-Generalized Born Surface Area) simulation were performed to evaluate the binding free energy, following the 100 (ns) production MD simulation in the periodic boundary condition (PBC) for the selected docked inhibitors with NS3. The residue-wise decomposition energy determined in our study revealed the active site region made the high stability with the potential inhibitors.

Machine learning the electric field response of condensed phase systems using perturbed neural network potentials

The interaction of condensed phase systems with external electric fields is of major importance in a myriad of processes in nature and technology, ranging from the field-directed motion of cells (galvanotaxis), to geochemistry and the formation of ice phases on planets, to field-directed chemical catalysis and energy storage and conversion systems including supercapacitors, batteries and solar cells. Molecular simulation in the presence of electric fields would give important atomistic insight into these processes but applications of the most accurate methods such as ab-initio molecular dynamics (AIMD) are limited in scope by their computational expense. Here we introduce Perturbed Neural Network Potential Molecular Dynamics (PNNP MD) to push back the accessible time and length scales of such simulations. We demonstrate that important dielectric properties of liquid water including the field-induced relaxation dynamics, the dielectric constant and the field-dependent IR spectrum can be machine learned up to surprisingly high field strengths of about 0.2 V Å−1 without loss in accuracy when compared to ab-initio molecular dynamics. This is remarkable because, in contrast to most previous approaches, the two neural networks on which PNNP MD is based are exclusively trained on molecular configurations sampled from zero-field MD simulations, demonstrating that the networks not only interpolate but also reliably extrapolate the field response. PNNP MD is based on rigorous theory yet it is simple, general, modular, and systematically improvable allowing us to obtain atomistic insight into the interaction of a wide range of condensed phase systems with external electric fields.

Synthesis, antimicrobial activity, molecular docking and MD simulation studies, in silico ADME investigations and thermo-acoustic studies of heterocyclic dihydropyrimidine substituted 1,3,4-thiadiazole scaffolds

The connections between the chemical structures and antibacterial properties of different functional groups including organic molecules were examined in this study. we have synthesized a novel series of 1,3,4-thiadiazole hybrids (5a-o) based on 3,4-dihydropyrimidines. After an effective synthesis, the compounds' structure was confirmed by using spectrum techniques like IR, 1H NMR, 13C NMR, and mass spectrometry. Elemental analyses were performed on a Perkin-Elmer 2400 CHN analyzer and the resulting data were within the accepted range (± 0.40) of the calculated values. The in vitro antimicrobial activity of the compounds against a range of bacterial and fungal species. Compound 5i showed potent inhibitory effects against P. aeruginosa with MIC values of 12.5 μg/ml S. aureus and A.niger were positively affected by compound 5k with MIC values of 12.5 μg/ml and 50 μg/ml, respectively. Based on the results, compounds 5a and 5o have moderate activity against C.albicans and E. coli with MIC values of 50 μg/ml and 62.5 μg/ml respectively. Furthermore, molecular docking and molecular dynamics simulations were used to identify the most stable conformation of newly discovered inhibitors. Thermo-acoustical properties of derivatives 5k and 5l in DMSO and DMF solvent have been determined at three different atmospheric pressure temperatures (298.15, 308.15, and 318.15)K. The components of the liquid mixture's molecular interactions have been taken into account to explain these results. The acquired results are interpreted in terms of interactions between solutes and solvents, providing insight into the compounds' potential to form or disrupt structures in DMSO and DMF solutions.

Targeting FAK, VEGF, and MTA1 proteins with Terminalia elliptica: a computational approach for anticancer activity.

Cancer remains a significant global health challenge, prompting exploration into alternative treatments such as those derived from natural compounds found in traditional medicine. Recent research has underscored the role of proteins like Focal Adhesion Kinase (FAK), Vascular Endothelial Growth Factor (VEGF), and Metastasis-Associated Protein 1 (MTA1) in driving cancer cell proliferation and survival. Here, we investigated the potential of a single molecule to modulate these key proteins involved in metastasis, offering a promising avenue for cancer therapy. Terminalia elliptica, commonly known as Asna, possesses a diverse range of medicinal properties, including antimicrobial, anti-inflammatory, anticancer, antidiabetic, antiaging, hepatoprotective, antioxidant, and neuroprotective activities. Our study aimed to explore the anticancer potential of Terminalia elliptica by identifying bioactive compounds capable of targeting FAK, VEGF, and MTA1 to impede cancer metastasis. Through in silico analysis, we conducted network analysis using Cytoscape to assess the significance of these bioactive compounds in the inhibition of signaling pathways driving metastasis. The utilization of these bioactives as potential candidates for targeted therapy of VEGF, FAK, and MTA1 regulated pathways was preliminarily assessed by Molecular Docking and MD Simulation. Our findings revealed that phytobioactives namely, Chebulinic Acid of Terminalia elliptica, exhibited notable binding affinity and interaction with FAK, and Chebulagic Acid with VEGF, and MTA1. This discovery holds promise as a novel therapeutic approach for combating cancer, offering potential benefits in cancer treatment and management.

PLS‐guided Mutant Recombination to Improve the Stability of Bovine Enterokinases Obtained by Directed Evolution

Activity and thermostability are critical yet challenging to improve simultaneously in enzymes. Using directed evolution, we previously identified bovine enterokinase (EKL) variants with enhanced soluble expression and thermal stability. Partial least squares (PLS) analysis of 321 EKL variants revealed the impact of individual mutations and identified neutral or detrimental mutations in top‐performing variants. Leveraging PLS rankings, we created new variants with fewer mutations and enhanced stability. Most original and PLS‐guided variants exhibited an activity‐stability trade‐off. However, two new triple‐ and quadruple‐mutants improved both activity and stability, surpassing the trade‐off limit. Recombining PLS‐guided mutations likely eliminated neutral or harmful mutations, enhancing stability. MD simulations linked residue‐specific dynamics with stability, pinpointing critical structural regions near aggregation‐prone areas. Our findings validate PLS as a potent strategy to enhance enzyme properties, complementing directed evolution.

Trp-cage-1,1,1,3,3,3-Hexafluoro-2-propanol-Water Interactions in a Nanosphere at 298 K.

MD simulations of the peptide Trp-cage dissolved in a solvent composed of 28% 1,1,1,3,3,3-hexafluoroisopropanol-water and contained within a nanosphere of 4.2 nm radius are described. To provide a thermal buffer, the nanosphere is submerged in a collection of liquid neopentane molecules, modeled as united atoms. It was found that the HFIP-water mixture demixes when confined under these conditions, with most of the fluoroalcohol becoming strongly associated with the walls of the nanosphere. The remaining HFIP interacts with the peptide and itself in the interior of the nanoshell. Diffusion of HFIP molecules near the surface of the nanoshell is limited, taking place over a small volume, while diffusion of the fluoroalcohol in the center of the nanoshell is more wide-ranging. By contrast, diffusion of the water in the solvent mixture appears to take place equivalently throughout the shell. The conformation of the peptide, distribution of solvent components around it, and the number and duration of the contacts with solvent molecules are calculated and contrasted to results previously reported for the bulk (non-nanocontained) system.