Protein-protein Docking Simulations Research Articles

Comparative Molecular Docking of Apigenin and Luteolin versus Conventional Ligands for TP-53, pRb, APOBEC3H, and HPV-16 E6: Potential Clinical Applications in Preventing Gynecological Malignancies.

This study presents a comparative analysis of molecular docking data, focusing on the binding interactions of the natural compounds apigenin and luteolin with the proteins TP-53, pRb, and APOBEC, in comparison to conventional pharmacological ligands. Advanced bioinformatics techniques were employed to evaluate and contrast binding energies, showing that apigenin and luteolin demonstrate significantly higher affinities for TP-53, pRb, and APOBEC, with binding energies of -6.9 kcal/mol and -6.6 kcal/mol, respectively. These values suggest strong potential for therapeutic intervention against HPV-16. Conventional ligands, by comparison, exhibited lower affinities, with energies ranging from -4.5 to -5.5 kcal/mol. Additionally, protein-protein docking simulations were performed to assess the interaction between HPV-16 E6 oncoprotein and tumor suppressors TP-53 and pRb, which revealed high binding energies around -976.7 kcal/mol, indicative of their complex interaction. A conversion formula was applied to translate these protein-protein interaction energies to a comparable scale for non-protein interactions, further underscoring the superior binding potential of apigenin and luteolin. These findings highlight the therapeutic promise of these natural compounds in preventing HPV-16-induced oncogenesis, warranting further experimental validation for clinical applications.

Unraveling modulation effects on albumin synthesis and inflammation by Striatin, a bioactive protein fraction isolated from Channa striata: In silico proteomics and in vitro approaches

Hypoalbuminemia, associated with inflammation in severely ill patients, can emerge due to decreased albumin production. Transforming growth factor-beta (TGF-β) and nuclear factor-kappa B (NF-κB) are critical signaling pathways responsible for decreased albumin expression. This study explores the protein content and modulation effects of Striatin on albumin synthesis and inflammation, employing in silico proteomics and in vitro investigations. In the in silico proteomics realm, LC/MS-MS protein sequencing, 3D modeling, protein-protein docking simulations, 100 ns molecular dynamics (MD) simulations, and MM/PBSA binding free energy calculations were carried out. Complementing this, in vitro studies examined Albumin gene expression and extracellular secretion in HepG2 cells subjected to lipopolysaccharides-induced hypoalbuminemia. Furthermore, the study probed Striatin's influence on the NF-ᴋB expression, given albumin's role as a negative acute-phase protein. The results unveiled nucleoside diphosphate kinase (NdK) and parvalbumin (PV) as the prominent constituents within Striatin. Notably, NdK and PV exhibited the ability to disrupt hydrogen bonds with specific residues in both TGF-β and NF-κB complexes, thereby enhancing their flexibility, akin to the action of the FKBP12 complex (antagonist complex). In the in vitro experiments, Striatin demonstrated a dose and time-dependent inhibition of hypoalbuminemia, with peak efficacy observed at a concentration of 20 μg/mL. At this concentration, Striatin also suppressed NF-κB expression when co-incubated with lipopolysaccharides. While these findings suggest potential inhibitory effects of Striatin on TGF-β and NF-κB activities, they are preliminary and warrant further investigation. This study highlights Striatin's potential as a therapeutic agent for inflammation-related hypoalbuminemia, though additional research is needed to fully validate these results.

Investigating the Mechanisms of Antibody Binding to Alpha-Synuclein for the Treatment of Parkinson's Disease.

Parkinson's disease (PD) is an idiopathic neurodegenerative disorder with the second-highest prevalence rate behind Alzheimer's disease. The pathophysiological hallmarks of PD are both degeneration of dopaminergic neurons in the substantia nigra pars compacta and the inclusion of misfolded α-synuclein (α-syn) aggregates known as Lewy bodies. Despite decades of research for potential PD treatments, none have been developed, and developing new therapeutic agents is a time-consuming and expensive process. Computational methods can be used to investigate the properties of drug candidates currently undergoing clinical trials to determine their theoretical efficiency at targeting α-syn. Monoclonal antibodies (mAbs) are biological drugs with high specificity, and Prasinezumab (PRX002) is an mAb currently in Phase II, which targets the C-terminus (AA 118-126) of α-syn. We utilized BioLuminate and PyMol for the structure prediction and preparation of the fragment antigen-binding (Fab) region of PRX002 and 34 different conformations of α-syn. Protein-protein docking simulations were performed using PIPER, and 3 of the docking poses were selected based on the best fit. Molecular dynamics simulations were conducted on the docked protein structures in triplicate for 1000 ns, and hydrogen bonds and electrostatic and hydrophobic interactions were analyzed using MDAnalysis to determine which residues were interacting and how often. Hydrogen bonds were shown to form frequently between the HCDR2 region of PRX002 and α-syn. Free energy was calculated to determine the binding affinity. The predicted binding affinity shows a strong antibody-antigen attraction between PRX002 and α-syn. RMSD was calculated to determine the conformational change of these regions throughout the simulation. The mAb's developability was determined using computational screening methods. Our results demonstrate the efficiency and developability of this therapeutic agent.

In silico-driven identification and experimental confirmation of antifungal proteins (AFPs) against Candidaalbicans

Mycoses infect millions of people annually across the world. The most common mycosis agent, Candida albicans is responsible for a great deal of illness and death. C. albicans infection is becoming more widespread and the current antifungals polyenes, triazoles, and echinocandins are less efficient against it. Investigating antifungal peptides (AFPs) as therapeutic is gaining momentum. Therefore, we used MALDI-TOF/MS analysis to identify AFPs and protein-protein docking to analyze their interactions with the C. albicans target protein. Some microorganisms with strong antifungal action against C. albicans were selected for the isolation of AFPs. Using MALDI-TOF/MS, we identified 3 AFPs Chitin binding protein (ACW83017.1; Bacillus licheniformis), the bifunctional protein GlmU (BBQ13478.1; Stenotrophomonas maltophilia), and zinc metalloproteinase aureolysin (BBA25172.1; Staphylococcus aureus). These AFPs showed robust interactions with C. albicans target protein Sap5. We deciphered some important residues in identified APFs and highlighted interaction with Sap5 through hydrogen bonds, protein-protein interactions, and salt bridges using protein-protein docking and MD simulations. The three discovered AFPs-Sap5 complexes exhibit different levels of stability, as seen by the RMSD analysis and interaction patterns. Among protein-protein interactions, the remarkable stability of the BBQ25172.1-2QZX complex highlights the role of salt bridges and hydrogen bonds. Identified AFPs could be further studied for developing successful antifungal candidates and peptide-based new antifungal therapeutic strategies as fresh insights into addressing antifungal resistance also.

Deciphering the genetic impact of signal peptide missense CTLA-4 polymorphism with rheumatoid arthritis in the Indian population: A case-control and in silico studies

Cytoplasmic T Lymphocyte Antigen-4 (CTLA-4) gene encodes for a glycoprotein, expressed on activated T-cells to transfer an inhibitory signal to control T-cell activation and proliferation. Techniques coupled with Real-time Polymerase Chain Reaction (PCR) and High-Resolution Melting Analysis (HRMA) were used to screen a missense signal peptide polymorphism (CTLA-4 + 49 A/G rs231775) in the Indian population to detect its association with Rheumatoid Arthritis (RA). Further, the resulting outcome was confirmed by Sanger’s sequencing technique, and genotype frequencies were calculated. In eukaryotic cells, the M domain of the Signal Recognition Particle (SRP-54) recognizes the N-terminal region of the Signal Peptide (SP) sequence. SP directs the polypeptide chain into the Sec-61 translocon of the Endoplasmic Reticulum (ER) for further protein modification. As the Single Nucleotide Polymorphism (SNP) rs231775 lies in the signal peptide region of CTLA-4, an in-silico study was also performed to predict the mRNA stability and SP-SRP protein interaction. From the study, it was observed that the genotype frequency of rs231775 SNP G/G homozygous dominant was significantly higher in RA patients than G/A heterozygous dominant and A/A homozygous recessive conditions (Odd Ratio (OR) = 2.0862; 95 % Confidence Interval (C.I) = 1.2584 to 3.4584; Relative Risk (RR) = 1.8507; p = 0.0044). Moreover, the rs231775 SNP G allele frequency was higher in RA than the control group G = 0.407 (40.7 %) vs 0.32 (32 %). In silico approaches of Protein-Protein docking and Molecular Dynamics (MD) simulation reveal CTLA-4 rs231775 SNP (G allele) has destabilized the SP-SRP protein complex, which may affect the translocation of CTLA-4 nascent polypeptide chains into the ER via activating Regulation of Aberrant Protein Production (RAPP) pathway.

Development of a subunit vaccine against the cholangiocarcinoma causing Opisthorchis viverrini: a computational approach.

Opisthorchis viverrini is the etiological agent of the disease opisthorchiasis and related cholangiocarcinoma (CCA). It infects fish-eating mammals and more than 10 million people in Southeast Asia suffered from opisthorchiasis with a high fatality rate. The only effective drug against this parasite is Praziquantel, which has significant side effects. Due to the lack of appropriate treatment options and the high death rate, there is a dire need to develop novel therapies against this pathogen. In this study, we designed a multi-epitope chimeric vaccine design against O. viverrini by using immunoinformatics approaches. Non-allergenic and immunogenic MHC-1, MHC-2, and B cell epitopes of three candidate proteins thioredoxin peroxidase (Ov-TPx-1), cathepsin F1 (Ov-CF-1) and calreticulin (Ov-CALR) of O. viverrini, were predicted to construct a potent multiepitope vaccine. The coverage of the HLA-alleles of these selected epitopes was determined globally. Four vaccine constructs made by different adjuvants and linkers were evaluated in the context of their physicochemical properties, antigenicity, and allergenicity. Protein-protein docking and MD simulation found that vaccines 3 was more stable and had a higher binding affinity for TLR2 and TLR4 immune receptors. In-silico restriction cloning of vaccine model led to the formation of plasmid constructs for expression in a suitable host. Finally, the immune simulation showed strong immunological reactions to the engineered vaccine. These findings suggest that the final vaccine construct has the potential to be validated by in vivo and in vitro experiments to confirm its efficacy against the CCA causing O. viverrini.

Citrullinating enzyme PADI4 and transcriptional repressor RING1B bind in cancer cells

Polycomb groups (PcGs) are transcriptional repressors, formed by a complex of several proteins, involved in multicellular development and cancer epigenetics. One of these proteins is the E3 ubiquitin-protein ligase RING1 (or RING1B), associated with the regulation of transcriptional repression and responsible for monoubiquitylation of the histone H2A. On the other hand, PADI4 is one of the human isoforms of a family of enzymes implicated in the conversion of arginine to citrulline, and it is also involved in the development of glioblastoma, among other types of cancers. In this work, we showed the association of PADI4 and RING1B in the nucleus and cytosol in several cancer cell lines by using immunofluorescence and proximity ligation assays. Furthermore, we demonstrated that binding was hampered in the presence of GSK484, an enzymatic PADI4 inhibitor, suggesting that RING1B could bind to the active site of PADI4, as confirmed by protein-protein docking simulations. In vitro and in silico findings showed that binding to PADI4 occurred for the isolated fragments corresponding to both the N-terminal (residues 1–221) and C-terminal (residues 228–336) regions of RING1B. Binding to PADI4 was also hampered by GSK484, as shown by isothermal titration calorimetry (ITC) experiments for the sole N-terminal region, and by both NMR and ITC for the C-terminal one. The dissociation constants between PADI4 and any of the two isolated RING1B fragments were in the low micromolar range (~2–10 μM), as measured by fluorescence and ITC. The interaction between RING1B and PADI4 might imply citrullination of the former, leading to several biological consequences, as well as being of potential therapeutic relevance for improving cancer treatment with the generation of new antigens.

Cracked actin filaments as mechanosensitive receptors

Actin filament networks are exposed to mechanical stimuli, but the effect of strain on actin filament structure has not been well established in molecular detail. This is a critical gap in understanding because the activity of a variety of actin-binding proteins has recently been determined to be altered by actin filament strain. We therefore used all-atom molecular dynamics simulations to apply tensile strains to actin filaments and find that changes in actin subunit organization are minimal in mechanically strained, but intact, actin filaments. However, a conformational change disrupts the critical D-loop to W-loop connection between longitudinal neighboring subunits, which leads to a metastable cracked conformation of the actin filament whereby one protofilament is broken prior to filament severing. We propose that the metastable crack presents a force-activated binding site for actin regulatory factors that specifically associate with strained actin filaments. Through protein-protein docking simulations, we find that 43 evolutionarily diverse members of the dual zinc-finger-containing LIM-domain family, which localize to mechanically strained actin filaments, recognize two binding sites exposed at the cracked interface. Furthermore, through its interactions with the crack, LIM domains increase the length of time damaged filaments remain stable. Our findings propose a new molecular model for mechanosensitive binding to actin filaments.

Enterocin DD14 can inhibit the infection of eukaryotic cells with enveloped viruses.

Bacteriocins are ribosomally synthesized bacterial peptides endowed with antibacterial, antiprotozoal, anticancer and antiviral activities. In the present study, we evaluated the antiviral activities of two bacteriocins, enterocin DD14 (EntDD14) and lacticaseicin 30, against herpes simplex virus type 1 (HSV-1), human coronavirus 229E (HCoV-229E) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Vero, Huh7 and Vero E6 cells, respectively. In addition, the interactions of these bacteriocins with the envelope glycoprotein D of HSV-1 and the receptor binding domains of HCoV-229E and SARS-CoV-2 have been computationally evaluated using protein-protein docking and molecular dynamics simulations. HSV-1 replication in Vero cells was inhibited by EntDD14 and, to a lesser extent, by lacticaseicin 30 added to cells after virus inoculation. EntDD14 and lacticaseicin 30 had no apparent antiviral activity against HCoV-229E; however, EntDD14 was able to inhibit SARS-CoV-2 in Vero E6 cells. Further studies are needed to elucidate the antiviral mechanism of these bacteriocins.

Targeted drug delivery to the thrombus by fusing streptokinase with a fibrin-binding peptide (CREKA): an in silico study.

Aim: Streptokinase has poor selectivity and provokes the immune response. In this study, we used in silico studies to design a fusion protein to achieve targeted delivery to the thrombus. Materials & methods: Streptokinase was analyzed computationally for mapping. The fusion protein modeling and quality assessment were carried out on several servers. The enzymatic activity and the stability of the fusion protein and its complex with plasminogen were assessed through molecular docking analysis and molecular dynamics simulation respectively. Results: Physicochemical properties analysis, protein quality assessments, protein-protein docking and molecular dynamics simulations predicted that the designed fusion protein is functionally active. Conclusion: Our results showed that this fusion protein might be a prospective candidate as a novel thrombolytic agent with better selectivity.

SurfPro-NN: A 3D point cloud neural network for the scoring of protein–protein docking models based on surfaces features and protein language models

Protein–protein interactions (PPI) play a crucial role in numerous key biological processes, and the structure of protein complexes provides valuable clues for in-depth exploration of molecular-level biological processes. Protein–protein docking technology is widely used to simulate the spatial structure of proteins. However, there are still challenges in selecting candidate decoys that closely resemble the native structure from protein–protein docking simulations. In this study, we introduce a docking evaluation method based on three-dimensional point cloud neural networks named SurfPro-NN, which represents protein structures as point clouds and learns interaction information from protein interfaces by applying a point cloud neural network. With the continuous advancement of deep learning in the field of biology, a series of knowledge-rich pre-trained models have emerged. We incorporate protein surface representation models and language models into our approach, greatly enhancing feature representation capabilities and achieving superior performance in protein docking model scoring tasks. Through comprehensive testing on public datasets, we find that our method outperforms state-of-the-art deep learning approaches in protein–protein docking model scoring. Not only does it significantly improve performance, but it also greatly accelerates training speed. This study demonstrates the potential of our approach in addressing protein interaction assessment problems, providing strong support for future research and applications in the field of biology.

Binding affinity between coronavirus spike protein and human ACE2 receptor

Coronaviruses (CoVs) pose a major risk to global public health due to their ability to infect diverse animal species and potential for emergence in humans. The CoV spike protein mediates viral entry into the cell and plays a crucial role in determining the binding affinity to host cell receptors. With particular emphasis on α- and β-coronaviruses that infect humans and domestic animals, current research on CoV receptor use suggests that the exploitation of the angiotensin-converting enzyme 2 (ACE2) receptor poses a significant threat for viral emergence with pandemic potential. This review summarizes the approaches used to study binding interactions between CoV spike proteins and the human ACE2 (hACE2) receptor. Solid-phase enzyme immunoassays and cell binding assays allow qualitative assessment of binding but lack quantitative evaluation of affinity. Surface plasmon resonance, Bio-layer interferometry, and Microscale Thermophoresis on the other hand, provide accurate affinity measurement through equilibrium dissociation constants (KD). In silico modeling predicts affinity through binding structure modeling, protein-protein docking simulations, and binding energy calculations but reveals inconsistent results due to the lack of a standardized approach. Machine learning and deep learning models utilize simulated and experimental protein-protein interaction data to elucidate the critical residues associated with CoV binding affinity to hACE2. Further optimization and standardization of existing approaches for studying binding affinity could aid pandemic preparedness. Specifically, prioritizing surveillance of CoVs that can bind to human receptors stands to mitigate the risk of zoonotic spillover.

Therapeutic potential of earthworm-derived proteins: Targeting NEDD4 for cardiovascular disease intervention

Targeting neuronal precursor cell-expressed developmentally down-regulated 4 (NEDD4) offers a promising strategy for cardiovascular therapies. NEDD4, a ubiquitin ligase enzyme, is crucial in protein degradation and cellular signaling. Earthworms (Lumbricus genus) are noteworthy for their rich biochemical composition and pharmacological properties. This study investigated the interactions between Lumbricus-derived proteins and NEDD4 to identify potential cardiovascular therapeutic candidates. Using advanced computational techniques, including 3D structure modeling, protein-protein docking simulations, 100 ns molecular dynamics (MD) simulations, and Molecular mechanics/Poisson–Boltzmann surface area (MM/PBSA) calculations, we assessed the binding affinities and functional impacts of these proteins on NEDD4 activity. The findings indicated that Lumbricus-derived proteins such as Lumbrokinase, heat shock protein, and elongation factor 1 (EF-1)-alpha showed activities similar to standard antagonists in modulating NEDD4. These results aligned with previous studies showing the inhibitory effects of heat shock protein and EF-1 on NEDD4 ubiquitination and ligase activity. Additionally, MM/PBSA calculations revealed favorable binding free energies for these compounds, underscoring their therapeutic potential in cardiovascular diseases. In conclusion, this study highlighted the potential of Lumbricus-derived compounds in cardiovascular disease therapy via the NEDD4 pathway, warranting further biochemical and preclinical validation and exploring broader therapeutic applications.

Structural Analyses of An Epidermal Growth Factor Receptor-Specific Single-Chain Fragment Variable via An In Silico Approach.

Single-chain fragment variable (scFv) antibodies were previously constructed of variable light and heavy chains joined by a (Gly4-Ser) 3 linker. The linker was created using molecular modeling software as a loop structure. Here, we introduce a protocol forin silico analysis of a complete scFv antibody that interacts with the epidermal growth factor receptor (EGFR). The homology modeling, with Pyrx of protein-protein docking and molecular dynamic simulation of the interacting scFv antibody and EGFR First, the authors used a protein structure modeling program and Python for homology modeling, and the antibody scFv structure was modeled for homology. The investigators downloaded Pyrx software as a platform in the docking study. The Molecular dynamic simulation was run using modeling software. Results show that when the MD simulation was subjected to energy minimization, the protein model had the lowest binding energy (-5.4 kcal/M). In addition, the MD simulation in this study showed that the docked EGFR-scFv antibody was stable for 20-75 ns when the movement of the structure increased sharply to 7.2 Å. In conclusion, in silicoanalysiswas performed, and the molecular docking and molecular dynamics simulations of the scFv antibody proved the effectiveness of the designed immune-therapeutic drug scFv as a specific drug therapy for EGFR.

Regulation Mechanism of Electron Transfer Reaction from Cytochrome C to Cytochrome C Oxidase

The respiratory chain in mitochondria is quite essential for the energy generation in cells, effectively producing “the energy currency in living organisms”, ATP. The driving force for this system is a series of electron transfers form Complex I (NADH:quinone oxidoreductase) to Complex IV (Cytochrome c Oxidase; CcO) in inner mitochondria membrane, and these electron transfer reactions promote the proton pumping from the matrix to the intermembrane space in mitochondria, and, by use of the resulting proton concentration gradient across the membrane, Complex V (ATP synthase) produces ATP. These series of electron transfer reactions terminate at CcO to reduce molecular oxygen to water molecules. The electrons required for this four-electron reduction of molecular oxygen in CcO are donated from a small soluble electron transfer hemoprotein, Cytochrome c (Cyt c), and this electron transfer is also associated with the proton pumping for the ATP generation in the respiratory chain. To examine the molecular regulation mechanism of the electron transfer reaction to terminate the electron transport chain for four-electron reduction of molecular oxygen and proton pumping for the ATP generation, we kinetically followed the electron transfer reaction and analyzed the electron transfer reaction by the Michaelis-Menten equation. By utilizing the Cyt c mutants, we identified the amino acid residues to regulate the electron transfer reactions (Biochem. J., 2020, 477,1565). The kinetic analysis of the electron transfer reaction also revealed that the apparent rate for the electron transfer reaction determined by the steady-state kinetics is much slower than the intramolecular electron transfer rate in the Cyt c – CcO complex. On the other hand, we successfully determined the interaction site between Cyt c and CcO by using NMR (Proc. Natl. Acad. Sci., 2011, 108, 12271) and protein-protein docking simulation (J. Biol. Chem., 2016, 291, 15320). The Cyt c – CcO complex structure estimated from the NMR and protein-protein docking simulation clearly showed that the two redox centers, heme iron in Cyt c and CuA in subunit II of CcO, were located more than 20 Å apart, and the complexes exhibiting shorter redox center distances were energetically unstable. Combined with the results from kinetic and structural studies, we proposed a “conformationally gated electron transfer mechanism”, where the thermal fluctuation of the protein structure controls the electron transfer rate. However, direct experimental evidence that protein conformational fluctuations regulate electron transfer reactions has not been obtained. In this study, we focused on the viscosity of the protein solution as a factor to regulate the conformational fluctuation of the proteins, based on the assumption that high viscosity of the solution suppresses the conformational fluctuation. The electron transfer reaction from Cyt c to CcO was examined in the high viscosity solution and NMR relaxation measurements under high viscosity were also performed to identify the local structural fluctuation responsible for the regulation of the electron transfer rate. Together with the transient complex structures between Cyt c and CcO estimated by the MD simulation, the regulation mechanism of the electron transfer from Cyt c to CcO based on the “conformational gating” will be discussed.

Crystal structures of glycoprotein D of equine alphaherpesviruses reveal potential binding sites to the entry receptor MHC-I.

Cell entry of most alphaherpesviruses is mediated by the binding of glycoprotein D (gD) to different cell surface receptors. Equine herpesvirus type 1 (EHV-1) and EHV-4 gDs interact with equine major histocompatibility complex I (MHC-I) to initiate entry into equine cells. We have characterized the gD-MHC-I interaction by solving the crystal structures of EHV-1 and EHV-4 gDs (gD1, gD4), performing protein-protein docking simulations, surface plasmon resonance (SPR) analysis, and biological assays. The structures of gD1 and gD4 revealed the existence of a common V-set immunoglobulin-like (IgV-like) core comparable to those of other gD homologs. Molecular modeling yielded plausible binding hypotheses and identified key residues (F213 and D261) that are important for virus binding. Altering the key residues resulted in impaired virus growth in cells, which highlights the important role of these residues in the gD-MHC-I interaction. Taken together, our results add to our understanding of the initial herpesvirus-cell interactions and will contribute to the targeted design of antiviral drugs and vaccine development.

Structural insights into the effect of mutations in the spike protein of SARS-CoV-2 on the binding with human furin protein

The SARS COV-2 and its variants are spreading around the world at an alarming speed, due to its higher transmissibility and the conformational changes caused by mutations. The resulting COVID-19 pandemic has imposed severe health consequences on human health. Several countries of the world including Pakistan have studied its genome extensively and provided productive findings. In the current study, the mCSM, DynaMut2, and I-Mutant servers were used to analyze the effect of identified mutations on the structural stability of spike protein however, the molecular docking and simulations approaches were used to evaluate the dynamics of the bonding network between the wild-type and mutant spike proteins with furin. We addressed the mutational modifications that have occurred in the spike protein of SARS-COV-2 that were found in 215 Pakistani's isolates of COVID-19 patients to study the influence of mutations on the stability of the protein and its interaction with the host cell. We found 7 single amino acid substitute mutations in various domains that reside in spike protein. The H49Y, N74K, G181V, and G446V were found in the S1 domain while the D614A, V622F, and Q677H mutations were found in the central helices of the spike protein. Based on the observation, G181V, G446V, D614A, and V622F mutants were found highly destabilizing and responsible for structural perturbation. Protein-protein docking and molecular simulation analysis with that of furin have predicted that all the mutants enhanced the binding efficiency however, the V622F mutant has greatly altered the binding capacity which is further verified by the KD value (7.1 E−14) and therefore may enhance the spike protein cleavage by Furin and increase the rate of infectivity by SARS-CoV-2. On the other hand, the total binding energy for each complex was calculated which revealed −50.57 kcal/mol for the wild type, for G181V −52.69 kcal/mol, for G446V −56.44 kcal/mol, for D614A −59.78 kcal/mol while for V622F the TBE was calculated to be −85.84 kcal/mol. Overall, the current finding shows that these mutations have increased the binding of Furin for spike protein and shows that D614A and V622F have significant effects on the binding and infectivity.

Exploring the interaction between T-cell antigen receptor-related genes and MAPT or ACHE using integrated bioinformatics analysis.

Alzheimer's disease (AD) is a neurodegenerative disease that primarily occurs in elderly individuals with cognitive impairment. Although extracellular β-amyloid (Aβ) accumulation and tau protein hyperphosphorylation are considered to be leading causes of AD, the molecular mechanism of AD remains unknown. Therefore, in this study, we aimed to explore potential biomarkers of AD. Next-generation sequencing (NGS) datasets, GSE173955 and GSE203206, were collected from the Gene Expression Omnibus (GEO) database. Analysis of differentially expressed genes (DEGs), gene ontology (GO) functional enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, and protein-protein networks were performed to identify genes that are potentially associated with AD. Analysis of the DEG based protein-protein interaction (PPI) network using Cytoscape indicated that neuroinflammation and T-cell antigen receptor (TCR)-associated genes (LCK, ZAP70, and CD44) were the top three hub genes. Next, we validated these three hub genes in the AD database and utilized two machine learning models from different AD datasets (GSE15222) to observe their general relationship with AD. Analysis using the random forest classifier indicated that accuracy (78%) observed using the top three genes as inputs differed only slightly from that (84%) observed using all genes as inputs. Furthermore, another data set, GSE97760, which was analyzed using our novel eigenvalue decomposition method, indicated that the top three hub genes may be involved in tauopathies associated with AD, rather than Aβ pathology. In addition, protein-protein docking simulation revealed that the top hub genes could form stable binding sites with acetylcholinesterase (ACHE). This suggests a potential interaction between hub genes and ACHE, which plays an essential role in the development of anti-AD drug design. Overall, the findings of this study, which systematically analyzed several AD datasets, illustrated that LCK, ZAP70, and CD44 may be used as AD biomarkers. We also established a robust prediction model for classifying patients with AD.

Post-translational modifications optimize the ability of SARS-CoV-2 spike for effective interaction with host cell receptors.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein is the prime target for vaccines, diagnostics, and therapeutic antibodies against the virus. While anchored in the viral envelope, for effective virulence, the spike needs to maintain structural flexibility to recognize the host cell surface receptors and bind to them, a property that can heavily depend upon the dynamics of the unresolved domains, most prominently the stalk. Construction of the complete, membrane-bound spike model and the description of its dynamics are critical steps in understanding the inner working of this key element of the viral infection by SARS-CoV-2. Combining homology modeling, protein-protein docking, and molecular dynamics (MD) simulations, we have developed a full spike structure in a native membrane. Multimicrosecond MD simulations of this model, the longest known single trajectory of the full spike, reveal conformational dynamics employed by the protein to explore the surface of the host cell. In agreement with cryogenic electron microscopy (cryo-EM), three flexible hinges in the stalk allow for global conformational heterogeneity of spike in the fully glycosylated system mediated by glycan-glycan and glycan-lipid interactions. The dynamical range of the spike is considerably reduced in its nonglycosylated form, confining the area explored by the spike on the host cell surface. Furthermore, palmitoylation of the membrane domain amplifies the local curvature that may prime the fusion. We show that the identified hinge regions are highly conserved in SARS coronaviruses, highlighting their functional importance in enhancing viral infection, and thereby, provide points for discovery of alternative therapeutics against the virus.

Sodium pump regulation in health and disease.

The sodium-potassium ATPase (NKA) is the ion motive ATP-dependent transporter that establishes Na+ and K+ gradients across the cell membrane, facilitating many physiological processes. In the heart, NKA activity is regulated by phospholemman (PLM), which PLM inhibits NKA by reducing its apparent affinity for Na+. In this study, we used time-correlated single-photon counting, FRET-microscopy, molecular dynamics simulations, and electrophysiology to investigate the stoichiometry of the native NKA-PLM regulatory complex. Our data suggest a stoichiometry of the NKA-PLM complex composed of two α subunits, two β subunits, and decorated with two PLM regulators. Protein-protein docking and molecular dynamics simulations generated a structural model of the regulatory complex. This model was characterized by contacts between cytoplasmic domains of α subunits, between β subunit extracellular domains, and between the highly-conserved α subunits’ M3 helix and the N-termini of opposing β subunit. The helix M3 shows significant movement during the pump's reaction cycle and mutations in M3 are associated with several diseases. Here, we studied the G301R mutant that is unable to correctly localize in the plasma membrane when expressed alone, but a milder G301A mutant localizes correctly. The G301R localization was improved in the presence of a WT pump suggesting a dominant negative effect of this mutation. Our simulations of the dimeric complexes revealed decreased affinity of the Na+ in position II, which is unbound during the simulated time window. Surprisingly, Na+ binding was not affected in the simulated WT or G301A/R monomers. Two-electrode voltage clamp experiments revealed decreased affinity of G301A for external Na+ and ion leak from G301R mutants supporting our simulation data. This combination of ex vivo and in silico experiments provides an insight into the quaternary structure of the NKA-PLM regulatory complex and helps to explain the mechanism of G301A/R mutation pathology.