Journal of Biomolecular Structure and Dynamics

The affinity maturation of Sars-Cov-1 VHH-72 nanobody from its germline predecessor has been studied at the molecular level. The effect of somatic mutations accumulated during affinity maturation process on flexibility, stability and affinity of the germline and affinity matured nanobody was studied. Affinity maturation results in loss of local flexibility in CDR of H3 and this resulted in a gain of affinity towards the antigen. Further affinity maturation was found to destabilize the nanobody. Mechanistically the loss of flexibility of the CDR H3 is due to the redistribution of hydrogen bond network due to somatic mutation A50T, also this contributes significantly to the destability of the nanobody. Unlike antibody, in nanobody the framework region is highly conserved and structural diversity in CDR is the determining factor in diverse antigen binding and also a factor contributing to the stability. This study provide insights into the interrelationship between flexibility, stability and affinity during affinity maturation in a nanobody.

CONSPECTUSO_ST_ABSAimsC_ST_ABSThe ribosomal protein (r-protein) of bacteria is composed of 2.6 MDa ribonucleoproteins of the 30S and 50S subunits, which are essential elements for protein translation. The translational initiation step is an intensive regulated multi-step reaction in protein biosynthesis. During bacterial protein synthesis, the correct reading frame of the mRNA defines when the initiator fMet-tRNAiMet binds to the start codon AUG at the P-site of the 30S subunit. The formation of the P-site of the 30S subunit initiation complex (30S-IC) is governed by three ubiquitous initiation factors (IFs) such as IF1, IF2, and IF3. IF2 protein is an essential player that plays during the last stage of the initiation process. Earlier, Stokes and his co-workers studied chemicals probes using 30K diverse drugs that induced cold-sensitive growth inhibition in the bacterium. The assay studies revealed, Lamotrigine (LTG) effectively binds at domain II of IF2 protein. In our research, we took an attempt in identifying promising active residues that could responsible for anti-bacterial bioactivity with help of computational studies. Computational MethodsIn the present study, initially, we performed C- backbone alignment with the retrieved IF2 chain from AlphaFold. Further, we utilized SiteMap and CastP for the identification of plausible active binding sites. Further, we bound LTG with the designated domain(s) of IF2 protein and studied its binding affinity potential with help of adaptive molecular dynamics simulations at atomic levels using Desmond. Key FindingsOur research findings have shown accurate results and we could able to prove the assertion in contrast with the findings of Stokes and his co-workers where the LTG bind at domain II of IF2 protein. The key interacting residue Glu179 was revealed to have strong hydrogen bonding contacts with LTG at the sub-nanomolar range. In addition, we predicted the alternative promising site I Further, we gained in-depth analysis for studying multiple sites, to understand the synergism inhibitory activity. Promisingly, LTG could be able to bound with at Site 1 showing better affinity over the proposed domain II and other predicted sites. The adaptive molecular dynamics studies confirmed the promising active residues SignificanceThe binding site predictions approach provides an insight for further development of anti-bacterial therapeutics that might helpful for bacteria disease management and exhibiting inhibitory activity against various strains.

p53 protein coded by the Tp53 gene is considered as one of the most intensively researched protein and mainly due to its role as a tumor suppressor, it acts as a tumor suppressor by carrying out two biologically complex processes namely Cell cycle arrest and apoptosis, In the oncogenic Y220C mutant p53, tyrosine is replaced by cysteine at 220th residue of the DNA binding Domain which causes the formation of a surface crevice, this specific mutation is responsible for approx. 100,000 cancer cases per year due to the destabilization and denaturation of the protein, as a result, the protein degrades at room temperature. In this work we carry out intensive Molecular Dynamic Simulations and Molecular Docking Studies to understand the structural dynamics of wild type p53 and changes the occurs in the mutant protein and also try to design lead against the druggable crevice and at the end of our study we used fragment-based optimization to come up with lead molecules which can act as scaffold for further drug development process

Cancer cell metabolism is commonly reprogrammed to favour glycolysis over oxidative phosphorylation, even under aerobic conditions, a phenomenon known as the Warburg effect. A key enzyme implicated in this shift is phosphoglycerate mutase 1 (PGAM1), which catalyses the conversion of 3-phosphoglycerate to 2-phosphoglycerate. The human enzyme is dependent on cofactor 2,3-bisphosphoglycerate which can phosphorylate and thereby activate the enzyme at histidine 11 (H11). A recently characterised moonlighting activity of pyruvate kinase M2 (PKM2), in its monomeric or dimeric forms, leads to phosphoenolpyruvate (PEP)-dependent phosphorylation of PGAM1 at the same position. Crucially, this phosphorylation is dependent on prior tyrosine 119 (Y119) phosphorylation of PGAM1 by Src kinase, itself activated by oncogenic growth factors. Using AlphaFold 3 (AF3), this study models the conformational changes induced by PGAM1 Y119 phosphorylation and investigates the molecular basis for its role in facilitating PEP-dependent phosphorylation at H11. Structural comparisons suggest that Y119 phosphorylation induces rearrangement of the C-terminal tail of PGAM1, opening the catalytic site around H11 to enable binding of phosphoenolpyruvate (PEP). Use of molecular docking (Webina, SwissDock, and DiffDock) found that AF3 generated models of PGAM1 with Y119 phosphorylation showed enhanced binding of PEP in comparison to non-phosphorylated PGAM1. However, extensive protein-protein docking (ClusPro, AF3 multimer generation) failed to identify configurations of PGAM1 and PKM2 where catalytic sites were proximal. Overall, this study supports a model in which phosphorylation of PGAM1 at Y119 enables access to the active site for PEP, thus enhancing its enzymatic activation. These findings underscore the critical role of post-translational modifications in modulating protein function and exemplify the utility of AF3 in predicting PTM-induced structural changes relevant to cancer metabolism.

Receptor-like kinases (RLKs) are plant proteins that form signaling circuits to transduce information through the plant cell membrane to the nucleus and activate processes that direct growth, development, stress response, and disease resistance. Upon sensing various environmental stress stimuli, RLKs interact with specific targets and recruits several other proteins to initiate the defense mechanism. Among many RLK subfamilies, leucine-rich repeat RLKs (LRR-RLKs) are the largest. Xa21, a member of LRR-RLK, is a vital receptor protein in rice plants that binds with bacterial RaxX21-sY, whereas OsSERK2 is a somatic embryogenic receptor kinase (SERK) acts as a coreceptor. This study focuses on the effect of a substitution mutation of aspartate128 with asparagine128 (D128N) in OsSERK2 on the interdependent binding pattern of the mentioned Xa21, RaxX21-sY, and OsSERK2 D128N proteins. The results showed that the D128N mutation in OsSERK2 can significantly change the interaction pattern of the critical residues of the OsSERK2 and affects its receptor-ligand (Xa21-RaxX21-sY) interaction in the complex.

H1N1, a subtype of influenza A virus, remains a global concern due to its ability to cause highly contagious, and fatal infections of the respiratory tract that can lead to seasonal pandemics. Factors such as genetic reassortment and antigenic shift due to the segmented nature of its RNA genome, lead to its rapid evolution that impacts drug and vaccine interactions. Thus, there is growing interest in identifying small-molecules against H1N1 to address the challenges posed by its ever-changing nature and enhance ability to combat the virus. By targeting the surface proteins crucial in the viral life cycle and interactions with host cells, we conducted structure-based virtual screening of 2,471 FDA-approved small molecules against H1N1s hemagglutinin (HA) and neuraminidase (NA) using molecular docking, and molecular dynamic simulations. A binding pocket for HA was identified at the interface of the three protomers, close to the fusion peptide, while in the case of NA the co-crystal ligand binding site was targeted. We identified 5 molecules, namely Econazole, Butoconazole, Miconazole, Isoconazole, and Tioconazole with higher binding affinity to HA and 4 molecules, namely Acarbose, Rutin, Paromomycin, and Idarubicin showing superior binding affinity to NA. Further molecular dynamic simulation of these molecules bound with HA and NA reaffirm the stability of the complexes. These molecules are known to have antifungal and antiviral properties. Thus, this study elucidates the importance of targeting HA and NA and paves the way for repurposing existing antivirals, antibacterials, and antifungals as inhibitors of the H1N1 viral entry into host cells.

Interaction of low-density lipoprotein receptors with proprotein convertase subtilisin/ kexin type 9 (PCSK9) plays a vital role in causing atherosclerosis. It is the hidden precursor of clinical myocardial infarction (MI), stroke, CVD and estimates 60% of deaths worldwide. The current need is to design small molecules to prevent the interaction between PCSK9 with LDL receptors. This study aims to evaluate the PCSK9 antagonistic effect of a derivative of Stevioside (also referred as Methylidene tetracyclo derivative) and atorvastatin. Also, a comparative study was performed to analyze the binding interaction of molecules inside the active and allosteric sites of PCSK9. The RCSB downloaded protein 7S5H and above said ligands were optimized to the local minima energy level and docked inside the active and allosteric sites. The stability of non-bonded interaction of complex was analyzed using Desmond MD simulation studies. The results of docking showed that the Methylidene tetracyclo molecule possesses a two-fold higher affinity of -10.159 kcal/mol in the active site and -10.824 kcal/mol in the allosteric site. The Phe377 amino acid made the Methylidene tetracyclo molecule orient inside the active site. Nine H-bonds with 5 amino acids of allosteric site increase the binding affinity compared to Atorvastatin. The MD simulation studies exposed that the nonbonded interaction of Methylidene tetracyclo molecule was stable throughout 100ns. This confirms the Methylidene tetracyclo molecule will be the better hit as well as the lead molecule to inhibit PCSK9 protein.

Malaria is a major global health issue due to the emergence of resistance to most of the available antimalarial drugs. There is an urgent need to discover new antimalarials to tackle the resistance issue. A CDK-like protein, Pfmrk from Plasmodium falciparum, plays a crucial role in regulating cell proliferation and shares 36.28% homology with humans CDK (hCDK7). Pfmrk complex with Pfcyc-1 and stimulates kinase activity. Also, Pfcyc-1 from P. falciparum, which has the highest sequence homology with human cyclin (Cyclin H), binds to and activates Pfmrk in a cyclin-dependent way. This is the first indication that cyclin subunits regulate human and plasmodial CDKs in a similar manner. In this study, molecular docking analysis of Pfmrk against the selected FDA-approved drugs acquired from the ZINC15 database. The top five drugs, Lurasidone, Vorapaxar, Donovex, Alvesco, and Orap, were screened based on binding energies of best-docked scores ranging between -8 kcal/mol and -12 kcal/mol. Based on Molecular dynamics simulations for 100ns, Lurasidone showed the highest binding affinity (-105.90 {+/-} 57.72 kJ/mol), followed by Donovex (-92.877 {+/-} 17.872 kJ/mol) and exhibited stable interactions with the amino acid residues present in the active site of Pfmrk. The outcomes of in silico investigation putatively suggested that Lurasidone and Donovex exhibit antimalarial potency and could be translated as potential Pfmrk inhibitors and developing new drugs based on further in-vitro studies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=130 SRC="FIGDIR/small/547694v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@1e14204org.highwire.dtl.DTLVardef@1b522e8org.highwire.dtl.DTLVardef@106df75org.highwire.dtl.DTLVardef@389899_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsWe investigated the potential FDA-approved drugs for repurposing against the modelled protein Pfmrk. Alvesco, Donovex, Lurasidone, Orap, and Vorapaxar are potential FDA-approved drugs firmly binding with Pfmrk. Molecular docking and simulation studies show that Donovex and Lurasidone are potential inhibitors of the modelled Pfmrk protein. Donovex and Lurasidone are potential drugs that act as kinase inhibitors by binding to the ATP -binding site of the enzyme Pfmrk.

In this paper, we used in silico analysis to shed light on the possible interaction between TMPRSS2 and SARS-CoV2 spike (S) protein by examining the role of TMPRSS2 single nucleotide polymorphisms (SNPs) in relation with susceptibility and inter-individual variability of SARS-CoV2 infection. First, we used molecular docking of human TMPRSS2 protein to predict the binding site of TMPRSS2, especially the TMPRSS2 link loops, in order to assess the effect TMPRSS2 SNPs. The latter lead to missense variants on the interaction between TMPRSS2 and SARS-CoV2 S protein. In a second step, we further refine our analysis by performing a structure-function analysis of the complexes using PyMol software, and finally by MD simulations to validate the as-obtained results. Our findings show that 17 SNPs among the 692 natural TMPRSS2 coding variants are in positions to influence the binding of TMPRSS2 with the viral S protein. All of them give more important interaction energy as assessed by docking. Among the 17 SNPs, four missense variants E389A, K392Q, T393S and Q438E lead to "directly increasing" the interaction affinity and 2 missense variants R470I and Y416C cause it "directly decreasing". The R470I and Y416C present in African and American population, respectively. While the other 4 SNP variants (E389A; K392Q; T393S and Q438E) are present only in the European population, which could link the viral infection susceptibility to demographic, geographic and genetic factors.

Parkinsons Disease (PD) is the second most common neurodegenerative disorder which creates devastating effects on the neurons of the mid brain. PARK2 and PARK6 genes encode the E3 ubiquitin ligase Parkin and serine/threonine protein kinase PINK1 (PTEN-induced kinase 1), respectively. Mutations in these proteins are responsible for majority of the early onset of PD. PINK1 protein along with Parkin are known to participate in the mitochondrial mitophagy pathway which selectively removes damaged mitochondria. Parkin with the help of its intra-domain interactions goes to an autoinhibited inactive state. Parkin ubiquitin activity is suppressed in this condition. Phosphorylation of the Serine65 residue, present in the Ubl domain of Parkin, is required for its ubiquitin ligase activity. PINK1 protein phosphorylates the Serine65 residue after its self-phosphorylation. After activation, Parkin protein ubiquitinates other damaged mitochondrial proteins for their degradation. Therefore, PINK1-Parkin interaction is important for the proper maintenance of the mitochondrial and neuronal fidelity. In this present work, we are trying to find out how the mutations in the PINK1 kinase domain would affect the modes of its interaction with the Ubl domain of Parkin. We used the three dimensional coordinates of the PINK1 and Ubl domain of Parkin from our previous work and selected the mutations P296L and G309D which have the abilities to hamper the said interactions between PINK1 and the Ubl domain of Parkin. We used molecular docking followed by molecular dynamics simulations to study the nature and structural dynamics of the binding interactions. The results from our study could provide an insight into the PINK1 mediated Parkin activation and plausible biochemical mechanism behind the onset of PD.

The interaction between Proprotein convertase subtilisin/kexin type 9 (PCSK9) and low-density lipoprotein receptors responsible for causing atherosclerosis. According to estimates, it causes 60% of fatalities worldwide and is the covert precursor to clinical myocardial infarction (MI), stroke, and CVD. Designing tiny compounds that inhibit PCSK9 from interacting with LDL receptors is the need of the hour. Through bioinformatics-based studies, this study seeks to assess the interactions between a derivative of tetrahydrocurcumin and PCSK9 Protein and compare them to interactions with the literature based studies of standard Atorvastatin. Additionally, comparison research was carried out to examine how the new compound interacts in the active and allosteric regions of PCSK9. The above-mentioned compound, a derivative of Tetrahydrocurcumin, was adjusted and optimized to the level of local minimum energy using the RCSBs downloaded PDB file 7S5H. By Desmond MD simulation studies, the stability of the non-bonded interactions of the complexes was examined. An affinity of -9.493 kcal/mol for the active site and -8.148 kcal/mol for the allosteric site was observed by docking studies in comparison with the standard molecule, atorvastatin. Also, the MMGBSA value of -50.7142 kcal/mol indicates the Tetrahydrocurcumin derivative binds well compare to the standard, atorvastatin. The Tetrahydro curcumin derivative molecule was able to orient into the active region with the help of Asp238, Thr377, and Ser381 amino acids. In comparison to atorvastatin, the binding affinity was raised by seven H-bonds with six amino acids and one {pi} interaction of Arg295 amino acids of the allosteric site. The Tetrahydro curcumin molecules nonbonded interaction was found to be stable for 100 ns by MD simulation tests. This demonstrates that the Tetrahydrocurcumin derivative molecule will prove to be an effective substrate to modify PCSK9 protein behavior.

The recent outbreak of severe acute respiratory syndrome (SARS) coronavirus (CoV)-2 (SARS-CoV-2) causing coronavirus disease (covid19) has posed a great threat to human health. Previous outbreaks of SARS-CoV and Middle East respiratory Syndrome CoV (MERS-CoV) from the same CoV family had posed similar threat to human health and economic growth. To date, not even a single drug specific to any of these CoVs has been developed nor any anti-viral vaccine is available for the treatment of diseases caused by CoVs. Subunits present in spike glycoproteins of SARS-CoV and SARS-CoV-2 are involved in binding to human ACE2 Receptor which is the primary method of viral invasion. As it has been observed in the previous studies that there are very minor differences in the spike glycoproteins of SARS-CoV and SARS-CoV-2. SARS-CoV-2 has an additional furin cleavage site that makes it different from SARS-CoV (Walls et al., 2020). In this study, we have analyzed spike glycoproteins of SARS-CoV-2 and SARS-CoV phylogenetically and subjected them to selection pressure analysis. Selection pressure analysis has revealed some important sites in SARS-CoV-2 and SARS-CoV spike glycoproteins that might be involved in their pathogenicity. Further, we have developed a potential multi-epitope vaccine candidate against SARS-CoV-2 by analyzing its interactions with HLA-B*15:03 subtype. This vaccine consists of multiple T-helper (TH) cells, B-cells, and Cytotoxic T-cells (CTL) epitopes joined by linkers and an adjuvant to increase its immunogenicity. Conservation of selected epitopes in SARS, MERS, and human hosts, suggests that the designed vaccine could provide cross-protection. The vaccine is designed in silico by following a reverse vaccinology method acknowledging its antigenicity, immunogenicity, toxicity, and allergenicity. The vaccine candidate that we have designed as a result of this work shows promising result indicating its potential capability of simulating an immune response.

Peptide-based drugs are widely used as therapeutic products that exhibit strong binding affinities with high specificity and low toxicity. To develop novel therapeutics against severe involuntary muscle movement and trembling disorder (Dyskinesia), the study explored the design of peptides that could target the nicotine acetylcholine receptor (nAChR) as antagonistic leads. The work aimed to suggest that the pharmacological interaction of snake venom toxins and its associated peptides with nAChRs could be exploited as plausible solutions for PD. Molecular docking and Molecular dynamics exercises were carried out using long chain neurotoxins (-BTx, 1NTN) to analyze the efficacies of their binding to nAChRs, with both the entire toxin and short peptide moieties. The study demonstrated that the novel-peptide (F8J2D7) system appears to possess better affinity with the nAChR than the entire toxin, which is a first step towards designing peptide-based drugs from snake venom proteins that could facilitate further research in the peptide-engineering and drug designing prospects for various neurodegenerative diseases.

The causative agent of gastroenteritis is Shiga toxin, which belongs to a functionally and structurally associated protein family despite each individual having a unique amino acid sequence. After entering the ER lumen and relocating the toxic domain to the cytoplasm, they alter the large subunit of rRNA, preventing protein synthesis and ribosomal damage. Shiga-like toxin-1 (SLT-1) subunit B targets glycolipid receptor Gb3, which plays a significant role in cytotoxicity. Though the mutational effect on subunit B is important for cytotoxicity study, we lack better understanding. Our present study targets the mutational impact of glycine protein at their 62th amino acid sequence of subunit B. For example, how it can alter the receptor-binding capacity and virulence. We used in silico method with GROMACS software suite (version 5.2, 2020.1) on Google Colab for a 100ns (100,000ps) simulation period and UCSF Chimera software for visualizing mutant and wild-type structure similarities. Surprisingly, RMSD, RMSF, and Rg trajectories from the simulation analysis indicated a more stable and compact mutant structure than the wild type. Principle component analysis (PCA) and SASA were visualized for the entire 100ns, which pointed towards homogeneity between both structures and more solvent accessibility in the mutant structure. This mutation may elevate receptor-binding and virulence capacity. Moreover, this finding can offer a better insight for future vaccine production.

Dihydroquinine (DHQ), is a quinine-based compound with anti-malarial properties. However, little is known about its mechanism of action against T. gondii inhibition, which shares similar biology with Plasmodium spp. In order to explore DHQ activity as an inhibitor of T. gondii using in vitro assays, we first used an in silico approach to decipher its mechanisms of action based on previous knowledge about its disruption of nucleic acid and protein synthesis. An in silico study was performed on T. gondii parasite replication, transcriptional and translational machinery to decipher the binding potentials of DHQ to some top selected enzymes. We report for the first time, using an in silico analysis that showed that DHQ binds strongly to DNA gyrase, Calcium Dependent Protein Kinase 1 (CDPK 1), and prolyl tRNA synthetase and thus could affect DNA replication, transcriptional and translational activities in T. gondii. Also, we found DHQ to effectively bind to mitochondria detoxifying enzymes (i.e., superoxide dismutase (SOD), peroxidoxin, and Catalase (CAT)). In conclusion, DHQ could be a lead compound for the treatment of toxoplasmosis when successfully evaluated using in vitro and in vivo models to confirm its effectiveness and safety.

Melioidosis, also known as Whitmores disease, is caused by the deadly pathogen Burkholderia pseudomallei and remains a significant global health concern, particularly in South Asia. The disease is contracted through exposure to contaminated soil, water, air, and food. Infected individuals often present with abscesses in internal organs such as the lungs, spleen, and liver, and in soft tissues, with severe cases leading to septic shock and acute pneumonia. The rising incidence and mortality rates, coupled with B. pseudomalleis ability to form biofilms and develop resistance to antibiotics like cephalosporins, make treatment increasingly challenging. This highlights the urgent need for novel therapeutic approaches. D-Alanine-D-Alanine ligase (Ddl), a crucial enzyme involved in the final stage of bacterial cell wall synthesis, which protects the pathogen from the hostile cellular environment of the host. While many bacteria have two isoforms of this enzyme, B. pseudomallei possesses only the DdlB isoform, presenting a significant vulnerability. Our study represents the first successful attempt to target DdlB through a combination of molecular docking and molecular dynamics simulations. These investigations provide strong evidence that Conivaptan acts as an effective inhibitor of DdlB, offering a novel therapeutic approach for combating melioidosis.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to mutate and evolve with the emergence of omicron (B.1.1.529) as the new variant of concern. The rapid spread of this variant regionally and globally could be an allusion to increased infectivity, transmissibility, and antibody resistance. The omicron variant has a large set of mutations in its spike protein, specifically in the receptor binding domain (RBD), reflecting their significance in ACE2 interaction and antibody recognition. We have carried out the present study to understand how these mutations structurally impact the binding of the antibodies to their target epitope. We have computationally evaluated the binding of different classes of RBD targeted antibodies, namely, CB6 (etesevimab), REGN10933 (casirivimab), S309 (sotrovimab), and S2X259 to the omicron mutation-induced RBD. Molecular dynamics simulations and binding free energy calculations unveil the binding affinity and stability of the antibody-RBD complexes. All the four antibodies show reduced binding affinity towards the omicron RBD. The therapeutic antibody CB6 aka etesevimab was substantially affected due to numerous omicron mutations occurring in its target epitope. This study provides a structural insight into the reduced efficacy of RBD targeting antibodies against the SARS-CoV-2 omicron variant.

The target of most cancer chemotherapeutic agents is to drive cancer cells toward death. A fine balance between anti-apoptotic and pro-apoptotic proteins is needed to maintain cellular homeostasis. Any shift favoring the pro-apoptotic ones is needed to drive cellular death in cancer chemotherapy. However, anti-apoptotic proteins such as Bcl-2 and Bcl-xL bind with pro-apoptotic proteins to hinder apoptosis mechanisms. Overexpression of these anti-apoptotic proteins lead to several cancers by preventing apoptosis. In this study, molecular docking, ADMET predictions, and molecular dynamics simulations were performed for the identification of potent inhibitors of anti-apoptotic Bcl-2 with compounds of Morus alba. Our study discovered that Quercetin-3-(6-Malonylglucoside) and Epigallocatechin gallate recorded excellent binding affinity with Bcl-2. Therefore, we conclude that compounds of Morus alba should be subjected to further experimental studies (in vitro and in vivo) in order to confirm the findings that they could be used as better options in cancer chemotherapy.

SARS-COV-2 is a pandemic virus causing COVID-19 disease which affects lungs and upper respiratory tract leading to progressive increase in the death rate worldwide. Currently, there are more than 123 million cases and over 2.71 million confirmed death caused by this virus. In this study, by utilizing an immunoinformatic approach, multiepitope-based vaccine is designed from the membrane protein which plays a vital role in the virion assembly of the novel-CoV. A total of 19 MHC class- I binders with HLA-A and HLA-B alleles have been selected with NetMHC pan EL 4.0 method from IEDB MHC-I prediction server. Four epitopes candidates from M-protein were selected based on the antigenicity, stability, immunogenicity, Ramachandran plot and scores with 100 % was taken for docking analysis with alleles HLA-A (PDB ID: 1B0R) and HLA-B (PDB ID: 3C9N) using ClusPro server. Among the four epitopes, the epitope FVLAAVYRI has the least binding energy and forms electrostatic, hydrogen and hydrophobic interactions with HLA-A (-932.8 Kcal/mol) and HLA-B (-860.7 Kcal/mol) which induce the T-cell response. Each HLA-A and HLA-B complex in the system environment achieves stable backbone configuration between 45-100 ns of MD simulation. This study reports a potent antigenic and immunogenic profile of FVLAAVYRI epitope from M-protein and further in vitro and in vivo validation is needed for its adaptive use as vaccine against COVID-19.

Chitin, comprising of repeating units of N-acetyl-glucosamine, is the second most abundant polymer occurring in wide range of insects, fungi, yeasts and plants. Chitinases hydrolyze chitin into chitooligomers which finds multifarious uses in various sectors and are gaining attention particularly as a biocontrol agent against chitin-containing insects and plant pathogens. Although fungi are a significant source of chitinases, the phylogenetic and functional diversity of the fungal enzymes is not well understood. In this study, we employed molecular modeling and simulation techniques to investigate the molecular characteristics of chitinase (PmChi) from a mycoparasitic fungus, Paraphaeosphaeria minitans, widely used as a biological control agent. The secondary structure of PmChi is predominantly random coil (51.24%), followed by alpha helices (28.44%) and extended strands (14.22%). PmChi contains the conserved chitinase sequence FDGLDIDWE at positions 178 to 186, where glutamic acid (E) acts as the catalytic proton donor, and aspartic acid (D) stabilizes the protein by accommodating substrate distortion. The protein surface of PmChi is rich in non-polar amino acid residues, while the active site contains more polar residues to facilitate the reaction. Key amino acids involved in catalytic activity include Trp146, Asp184, Glu186, Tyr187, Pro228, Met252, Tyr254, and Asp255. Molecular simulations demonstrated that PmChi maintained stability during interaction with chitotriose. Residual flexibility, hydrogen bonding, and structural packing showed consistent trajectories with no significant perturbations throughout the simulation. The free energy calculations for the PmChi-chitotriose complex indicated that MM/PBSA calculations are more accurate for analyzing enzyme-carbohydrate interactions. These findings enhance our understanding of the structural properties and functional dynamics of chitinase from P. minitans and provide a platform for future research and applications.