In silico approaches for the identification of potential allergens among hypothetical proteins from Alternaria alternata and its functional annotation.

Direct exposure to the fungal species Alternaria alternata is a major risk factor for the development of asthma, allergic rhinitis, and inflammation. As of November 23rd 2020, the NCBI protein database showed 11,227 proteins from A. alternata genome as hypothetical proteins (HPs). Allergens are the main causative of several life-threatening diseases, especially in fungal infections. Therefore, the main aim of the study is to identify the potentially allergenic inducible proteins from the HPs in A. alternata and their associated functional assignment for the complete understanding of the complex biological systems at the molecular level. AlgPred and Structural Database of Allergenic Proteins (SDAP) were used for the prediction of potential allergens from the HPs of A. alternata. While analyzing the proteome data, 29 potential allergens were predicted by AlgPred and further screening in SDAP confirmed the allergic response of 10 proteins. Extensive bioinformatics tools including protein family classification, sequence-function relationship, protein motif discovery, pathway interactions, and intrinsic features from the amino acid sequence were used to successfully predict the probable functions of the 10 HPs. The functions of the HPs are characterized as chitin-binding, ribosomal protein P1, thaumatin, glycosyl hydrolase, and NOB1 proteins. The subcellular localization and signal peptide prediction of these 10 proteins has further provided additional information on localization and function. The allergens prediction and functional annotation of the 10 proteins may facilitate a better understanding of the allergenic mechanism of A. alternata in asthma and other diseases. The functional domain level insights and predicted structural features of the allergenic proteins help to understand the pathogenesis and host immune tolerance. The outcomes of the study would aid in the development of specific drugs to combat A. alternata infections.

Study of an inhibitory effect of plant polyphenolic compounds against digestive enzymes using bench-working experimental evidence predicted by molecular docking and dynamics.

The substantial nutritional content and diversified biological activity of plant-based nutraceuticals are due to polyphenolic chemicals. These chemicals are important and well-studied plant secondary metabolites. Their protein interactions are extensively studied. This relationship is crucial for the logical development of functional food and for enhancing the availability and usefulness of polyphenols. This study highlights the influence of protein types and polyphenols on the interaction, where the chemical bindings predominantly consist of hydrophobic interactions and hydrogen bonds. The interaction between polyphenolic compounds (PCs) and digestive enzymes concerning their inhibitory activity has not been fully studied. Therefore, we have examined the interaction of four digestive enzymes (α-amylase, pepsin, trypsin, and α-chymotrypsin) with four PCs (curcumin, diosmin, morin, and 2',3',4'-trihydroxychalcone) through in silico and in vitro approaches. In vitro plate assays, enzyme kinetics, spectroscopic assays, molecular docking, and simulations were performed. We observed all these PCs have significant docking scores and preferable interaction with the active site of the digestive enzymes, resulting in the reduction of enzyme activity. The enzyme-substrate binding mechanism was determined using the Lineweaver Burk plot, indicating that the inhibition occurred competitively. Among four PCs diosmin and morin has the highest interaction energy over digestive enzymes with IC50 value of 1.13 ± 0.0047 and 1.086 ± 0.0131 µM. Kinetic studies show that selected PCs inhibited pepsin, trypsin, and chymotrypsin competitively and inhibited amylase in a non-competitive manner, especially by 2',3',4'-trihydroxychalcone. This study offers insights into the mechanisms by which the selected PCs inhibit the enzymes and has the potential to enhance the application of curcumin, diosmin, morin, and 2',3',4'-trihydroxychalcone as natural inhibitors of digestive enzymes.

Identification of potent anti-fibrinolytic compounds against plasminogen and tissue-type plasminogen activator employing in silico approaches.

The zymogen protease Plasminogen (Plg) and its active form plasmin (Plm) carry out important functions in the blood clot disintegration (breakdown of fibrin fibers) process. Inhibition of plasmin effectively reduces fibrinolysis to circumvent heavy bleeding. Currently, available Plm inhibitor tranexamic acid (TXA) used for treating severe hemorrhages is associated with an increased incidence of seizures which in turn were traced to gamma-aminobutyric acid antagonistic activity (GABAa) in addition to having multiple side effects. Fibrinolysis can be suppressed by targeting the three important protein domains: the kringle-2 domain of tissue plasminogen activator, the kringle-1 domain of plasminogen, and the serine protease domain of plasminogen. In the present study, one million molecules were screened from the ZINC database. These ligands were docked to their respective protein targets using Autodock Vina, Schrödinger Glide, and ParDOCK/BAPPL+. Thereafter, the drug-likeness properties of the ligands were evaluated using Discovery Studio 3.5. Subsequently, we subjected the protein-ligand complexes to molecular dynamics simulation of 200 ns in GROMACS. The identified ligands P76(ZINC09970930), C97(ZINC14888376), and U97(ZINC11839443) for each protein target are found to impart higher stability and greater compactness to the protein-ligand complexes. Principal component analysis (PCA) implicates, that the identified ligands occupy smaller phase space, form stable clusters, and provide greater rigidity to the protein-ligand complexes. Molecular Mechanics Poisson-Boltzmann Surface Area (MMPBSA) analysis reveals that P76, C97, and U97 exhibit better binding free energy (ΔG) when compared to that of the standard ligands. Thus, our findings can be useful for the development of promising anti-fibrinolytic agents.Communicated by Ramaswamy H. Sarma.

Structural and functional characterization of 6-phosphogluconate dehydrogenase in Plasmodium falciparum (3D7) and identification of its potent inhibitors.

The malarial parasite Plasmodium falciparum predominantly causes severe malaria and deaths worldwide. Moreover, resistance developed by P. falciparum to frontline drugs in recent years has markedly increased malaria-related deaths in South Asian Countries. Ribulose 5-phosphate and NADPH synthesized by Pentose Phosphate Pathway (PPP) act as a direct precursor for nucleotide synthesis and P. falciparum survival during oxidative challenges in the intra-erythrocytic growth phase . In the present study, we have elucidated the structure and functional characteristics of 6-phosphogluconate dehydrogenase (6PGD) in P. falciparum and have identified potent hits against 6PGD by pharmacophore-based virtual screening with ZINC and ChemBridge databases. Molecular docking and Molecular dynamics simulation, binding free energies (MMGBSA & MMPBSA), and Density Functional Theory (DFT) calculations were integratively employed to validate and prioritize the most potential hits. The 6PGD structure was found to have an open and closed conformation during MD simulation. The apo form of 6PGD was found to be in closed conformation, while a open conformation attributed to facilitating binding of cofactor. It was also inferred from the conformational analysis that the small domain of 6PGD has a high influence in altering the conformation that may aid in open/closed conformation of 6PGD. The top three hits identified using pharmacophore hypotheses were ChemBridge_11084819, ChemBridge_80178394, and ChemBridge_17912340. Though all three hits scored a high glide score, MMGBSA, and favorable ADMET properties, ChemBridge_11084819 and ChemBrdige_17912340 showed higher stability and binding free energy. Moreover, these hits also featured stable H-bond interactions with the active loop of 6PGD with binding free energy comparable to substrate-bound complex. Therefore, the ChemBridge_11084819 and ChemBridge_17912340 moieties demonstrate to have high therapeutic potential against 6PGD in P. falciparum.Communicated by Ramaswamy H. Sarma.

Discovery of plant-based phytochemical as effective antivirals that target the non-structural protein C of the Nipah virus through computational methods.

Nipah Virus (NiV) belongs to the Paramyxoviridae family and was first identified during an outbreak in Malaysia. Some initial symptoms include mild fever, headache and sore throat, which could escalate to respiratory illness and brain inflammation. The mortality rate of NiV infection can range from 40% to 75%, which is quite high. This is mainly due to the lack of efficient drugs and vaccines. In most instances, NiV is transmitted from animals to humans. Non-Structural Proteins (C, V and W) of the Nipah virus impede the host immune response by obstructive the JAK/STAT pathway. However, Non-Structural Proteins - C (NSP-C) plays a vital role in NiV pathogenesis, which includes IFN antagonist activity and viral RNA production. In the present study, the full-length structure of NiV-NSP-C was predicted using computational modelling, and the stability of the structure was analysed using 200 ns molecular dynamic (MD) simulation. Further, the structure-based virtual screening identified five potent phytochemicals (PubChem CID: 9896047, 5885, 117678, 14887603 and 5461026) with better binding affinity against NiV-NSP-C. DFT studies clearly showed that the phytochemicals had higher chemical reactivity, and the complex MD simulation depicted that the identified inhibitors exhibited stable binding with NiV-NSP-C. Furthermore, experimental validation of these identified phytochemicals would likely control the infection of NiV.Communicated by Ramaswamy H. Sarma.

Unveiling the anti-glioma potential of a marine derivative, Fucoidan: its synergistic cytotoxicity with Temozolomide-an in vitro and in silico experimental study.

Glioma coined as a "butterfly" tumor associated with a dismal prognosis. Marine algal compounds with the richest sources of bioactive components act as significant anti-tumor therapeutics. However, there is a paucity of studies conducted on Fucoidan to enhance the anti-glioma efficacy of Temozolomide. Therefore, the present study aimed to evaluate the synergistic anti-proliferative, anti-inflammatory and pro-apoptotic effects of Fucoidan with Temozolomide in in vitro and in silico experimental setup. The anti-proliferative effects of Temozolomide and Fucoidan were evaluated on C6 glioma cells by MTT and migration assay. Modulation of inflammatory markers and apoptosis induction was affirmed at the morphological and transcriptional level by dual staining and gene expression. Molecular docking (MD) and molecular dynamics simulation (MDS) studies were performed against the targets to rationalize the inhibitory effect. The dual-drug combination significantly reduced the cell viability and migration of glioma cells in a synergistic dose-dependent manner. At the molecular level, the dual-drug combination significantly down-regulated inflammatory genes with a concomitant upregulation of pro-apoptotic marker. In consensus with our in vitro findings, molecular docking and simulation studies revealed that the anti-tumor ligands: Temozolomide, Fucoidan with 5-(3-Methy1-trizeno)-imidazole-4-carboxamide (MTIC), and 4-amino-5-imidazole-carboxamide (AIC) had the potency to bind to the inflammatory proteins at their active sites, mediated by H-bonds and other non-covalent interactions. The dual-drug combinatorial treatment synergistically inhibited the proliferation, migration of glioma cells and promoted apoptosis; conversely with the down-regulation of inflammatory genes. However, pre-clinical experimental evidence is warranted for the possible translation of this combination. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03814-6.

Non-Structural Protein-W61 as a Novel Target in Severe Fever with Thrombocytopenia Syndrome Virus (SFTSV): An In-Vitro and In-Silico Study on Protein-Protein Interactions with Nucleoprotein and Viral Replication.

The non-structural protein (NSs) and nucleoprotein (NP) of the severe fever with thrombocytopenia syndrome virus (SFTSV) encoded by the S segment are crucial for viral pathogenesis. They reside in viroplasm-like structures (VLS), but their interaction and their significance in viral propagation remain unclear. Here, we investigated the significance of the association between NSs and NP during viral infection through in-silico and in-vitro analyses. Through in-silico analysis, three possible binding sites were predicted, at positions C6S (Cystein at 6th position to Serine), W61Y (Tryptophan 61st to Tyrosine), and S207T (Serine 207th to Threonine), three mutants of NSs were developed by site-directed mutagenesis and tested for NP interaction by co-immunoprecipitation. NSsW61Y failed to interact with the nucleoprotein, which was substantiated by the conformational changes observed in the structural analyses. Additionally, molecular docking analysis corroborated that the NSW61Y mutant protein does not interact well compared to wild-type NSs. Over-expression of wild-type NSs in HeLa cells increased the SFTSV replication by five folds, but NSsW61Y exhibited 1.9-folds less viral replication than wild-type. We demonstrated that the W61Y alteration was implicated in the reduction of NSs-NP interaction and viral replication. Thus, the present study identified a critical NSs site, which could be targeted for development of therapeutic regimens against SFTSV.

In silico analysis of Diosmetin as an effective chemopreventive agent against prostate cancer: molecular docking, validation, dynamic simulation and pharmacokinetic prediction-based studies.

Prostate cancer is the second most dangerous cancer type worldwide. While various treatment options are present i.e. agonists and antagonists, their utilization leads to adverse effects and due to this resistance developing, ultimately the outcome is remission. So, to overcome this issue, we have undertaken an in-silico investigation to identify promising and unique flavonoid candidates for combating prostate cancer. Using GOLD software, the study assessed the effectiveness of 560 natural secondary polyphenols against CDKN2. Protein Data Bank was used to retrieve the 3D crystal structure of CDKN2 (PDB Id: 4EK3) and we retrieved the structure of selected secondary polyphenols from the PubChem database. The compound Diosmetin shows the highest GOLD score with the selected Protein i.e. CDKN2 which is 58.72. To better understand the 2-dimensional and 3-dimensional interactions, the interacting amino acid residues were visualised using Discovery Studio 3.5 and Maestro 13.5. Using Schrodinger-Glide, the Diosmetin and CDKN2 were re-docked, and decoy ligands were docked to CDKN2, which was used to further ascertain the study. The ligands with the highest Gold score were forecasted for pharmacokinetics characteristics, and the results were tabulated and analysed. Utilising the Gromacs software and Desmond packages, 100 ns of Diosmetin molecular dynamics simulations were run to evaluate the structural persistence and variations of protein-ligand complexes. Additionally, our investigation revealed that Diosmetin had a better binding affinity with CDKN2 measuring 58.72, and it also showed remarkable stability across a 100-ns simulation. Thus, following in-vitro and in-vivo clinical studies, diosmetin might lead to the Prostate regimen.Communicated by Ramaswamy H. Sarma.

Discovery of potent inhibitors targeting Glutathione S-transferase of Wuchereria bancrofti: a step toward the development of effective anti-filariasis drugs.

Lymphatic filariasis (LF) is one of the major health problems for the human kind in developing countries including India. LF is caused by three major nematodes namely Wuchereria bancrofti, Brugia malayi, and Brugia timori. The recent statistics of World Health Organization (WHO) showed that 51 million people were affected and 863 million people from 47 countries around worldwide remain threatened by LF. Among them, 90% of the filarial infection was caused by the nematode W. bancrofti. Approved drugs were available for the treatment of LF but many of them developed drug resistance and no longer effective in all stages of the infection. In the current research work, we explored the Glutathione S-transferase (GST) of W. bancrofti, the key enzyme responsible for detoxification that catalyzes the conjugation of reduced GSH (glutathione) to xenobiotic compounds. Initially, we analyzed the stability of the WbGST through 200 ns MD simulation and further structure-based virtual screening approach was applied by targeting the substrate binding site to identify the potential leads from small molecule collection. The in silico ADMET profiles for the top-ranked hits were predicted and the predicted non-toxic lead molecules showed the highest docking score in the range of - 12.72 kcal/mol to - 11.97 kcal/mol. The cross docking of the identified hits with human GST revealed the potential binding specificity of the hits toward WbGST. Through WbGST-lead complex simulation, the lead molecules were observed to be stable and also intactly bound within the binding site of WbGST. Based on the computational results, the five predicted non-toxic molecules were selected for the in vitro assay. The molecules showed significant percentage of inhibition against the filarial worm Setaria digitata which is the commonly used model organism to evaluate the filarial activity. In addition, the molecules also showed better IC50 than the standard drug ivermectin. The identified lead molecules will lay a significant insight for the development of new drugs with higher specificity and lesser toxicity to control and treat filarial infections.

Triphenylphosphonium conjugated gold nanotriangles impact Pi3K/AKT pathway in breast cancer cells: a photodynamic therapy approach.

Although gold nanoparticles based photodynamic therapy (PDT) were reported to improve efficacy and specificity, the impact of surface charge in targeting cancer is still a challenge. Herein, we report gold nanotriangles (AuNTs) tuned with anionic and cationic surface charge conjugating triphenylphosphonium (TPP) targeting breast cancer cells with 5-aminoleuvinic acid (5-ALA) based PDT, in vitro. Optimized surface charge of AuNTs with and without TPP kill breast cancer cells. By combining, 5-ALA and PDT, the surface charge augmented AuNTs deliver improved cellular toxicity as revealed by MTT, fluorescent probes and flow cytometry. Further, the 5-ALA and PDT treatment in the presence of AuNTs impairs cell survival Pi3K/AKT signaling pathway causing mitochondrial dependent apoptosis. The cumulative findings demonstrate that, cationic AuNTs with TPP excel selective targeting of breast cancer cells in the presence of 5-ALA and PDT.

Computational identification and experimental validation of anti-filarial lead molecules targeting metal binding/substrate channel residues of Cu/Zn SOD1 from Wuchereria bancrofti.

Lymphatic filariasis (LF) is a neglected mosquito-borne parasitic disease, widely caused by Wuchereria bancrofti (Wb) in tropical and sub-tropical countries. During a blood meal, the filarial nematodes are transmitted to humans by the infected mosquito. To counter attack the invaded nematodes, the human immune system produces reactive oxygen species. However, the anti-oxidant enzymes of nematodes counteract the host oxidative cytotoxicity. Cu/Zn Superoxide dismutase (SOD1), a member of antioxidant enzymes and are widely used by the nematodes to sustain the host oxidative stress across its lifecycle, hence targeting SOD1 to develop suitable drug molecules would help to overcome the problems related to efficacy and activity of drugs upon different stages of nematodes. In order to find the potent inhibitors, a three-dimensional structure of Cu/Zn WbSOD1 was modelled and the structural stability was analysed through simulation studies. The structure-guided virtual screening approach has been used to identify lead molecules from the ChemBridge based on the docking score, ADMET properties and protein-ligand complex stability analysis. The identified compounds were observed to interact with the copper, metal binding residues (His48, His63, His80 and His120) and catalytically important residue Arg146, which play a crucial role in the disproportionation of incoming superoxide radicals of Cu/Zn WbSOD1. Further, in vitro validation of the selected leads in the filarial worm Setaria digitata exhibited higher inhibition and better IC50 compared to the standard drug ivermectin. Thus, the identified leads could potentially inhibit enzyme activity, which could subsequently act as drug candidates to control LF.Communicated by Ramaswamy H. Sarma.

Functional Characterization, Mechanism, and Mode of Action of Putative Streptomycin Adenylyltransferase from Serratia marcescens.

Nosocomial infections are serious threats to the entire world in healthcare settings. The major causative agents of nosocomial infections are bacterial pathogens, among which Enterobacteriaceae family member Serratia marcescens plays a crucial role. It is a gram-negative opportunistic pathogen, predominantly affecting patients in intensive-care units. The presence of intrinsic genes in S. marcescens led to the development of resistance to antibiotics for survival. Complete scanning of the proteome, including hypothetical and partially annotated proteins, paves the way for a better understanding of potential drug targets. The targeted protein expressed in E. coli BL21 (DE3) pLysS cells has shown complete resistance to aminoglycoside antibiotic streptomycin (>256 MCG). The recombinant protein was purified using affinity and size-exclusion chromatography and characterized using SDS-PAGE, western blotting, and MALDI-TOF analysis. Free phosphate bound to malachite green was detected at 620 nm, evident of the conversion of adenosine triphosphate to adenosine monophosphate during the adenylation process. Similarly, in the chromatographic assay, adenylated streptomycin absorbed at 260 nm in AKTA (FPLC), confirming the enzyme-catalyzed adenylation of streptomycin. Further, the adenylated product of streptomycin was confirmed through HPLC and mass spectrometry analysis. In conclusion, our characterization studies identified the partially annotated hypothetical protein as streptomycin adenylyltransferase.

Environmental impact assessment of algal bloom Noctiluca scintillans in the Mandapam group of Islands, Gulf of Mannar Biosphere Reserve, Southeast coast of India.

The present investigation accounts for the environmental impact assessment of an intense algal bloom caused by the dinoflagellate Noctilucas scintillans. The bloom was first observed on the 10th of September 2019, in the vicinity of the Mandapam group of Islands, spreading from Rameswaram Coast in the North (9° 14' 15″ N, 79° 9' 46″ E) to Hare Island in the South (9° 14' 51″ N, 79° 5' 48″ E). The coastal waters in and around the Mandapam region appeared dark green, and the microscopic examination of the water sample revealed the presence of N. scintillans in large numbers. N. scintillans is a bioluminescent organism; it is inflated and sub-spherical in shape, and the size of the organisms ranged from 350 to 1300 microns. During the intense periods of the bloom, the average density of N. scintillans was recorded with 226.5 × 103 cells/l, and the dissolved oxygen content was very low and the ammonia content was extremely high in certain sites (avg. 4.3635 µm/l). Intensive bloom may lead to a loss of biodiversity in the affected areas of the region. Subsequent investigations indicated that the resilience of the ecosystem in response to natural adversity.

The Updated Review on Plant Peptides and Their Applications in Human Health.

Biologically active plant peptides, consisting of secondary metabolites, are compounds (amino acids) utilized by plants in their defense arsenal. Enzymatic processes and metabolic pathways secrete these plant peptides. They are also known for their medicinal value and have been incorporated in therapeutics of major human diseases. Nevertheless, its limitations (low bioavailability, high cytotoxicity, poor absorption, low abundance, improper metabolism, etc.) have demanded a need to explore further and discover other new plant compounds that overcome these limitations. Keeping this in mind, therapeutic plant proteins can be excellent remedial substitutes for bodily affliction. A multitude of these peptides demonstrates anti-carcinogenic, anti-microbial, anti-HIV, and neuro-regulating properties. This article's main aim is to list out and report the status of various therapeutic plant peptides and their prospective status as peptide-based drugs for multiple diseases (infectious and non-infectious). The feasibility of these compounds in the imminent future has also been discussed.

A Novel Prototype Biosensor Array Electrode System for Detecting the Bacterial Pathogen Salmonella typhimurium.

Salmonellosis caused by Salmonella sp. has long been reported all over the world. Despite the availability of various diagnostic methods, easy and effective detection systems are still required. This report describes a dialysis membrane electrode interface disc with immobilized specific antibodies to capture antigenic Salmonella cells. The interaction of a specific Salmonella antigen with a mouse anti-Salmonella monoclonal antibody complexed to rabbit anti-mouse secondary antibody conjugated with HRP and the substrate o-aminophenol resulted in a response signal output current measured using two electrode systems (cadmium reference electrode and glassy carbon working electrode) and an agilent HP34401A 6.5 digital multimeter without a potentiostat or applied potential input. A maximum response signal output current was recorded for various concentrations of Salmonella viz., 3, 30, 300, 3000, 30,000 and 300,000 cells. The biosensor has a detection limit of three cells, which is very sensitive when compared with other detection sensors. Little non-specific response was observed using Streptococcus, Vibrio, and Pseudomonas sp. The maximum response signal output current for a dialysis membrane electrode interface disc was greater than that for gelatin, collagen, and agarose. The device and technique have a range of biological applications. This novel detection system has great potential for future development and application in surveillance for microbial pathogens.

Potential inhibitors for peroxiredoxin 6 of W. bancrofti: A combined study of modelling, structure-based drug design and MD simulation.

Lymphatic filariasis (LF), a mosquito-borne parasitic disease caused by nematode Wuchereria bancrofti in tropical and sub-tropical countries. These nematodes are transferred into the human host when the infected mosquito carrying L3 larvae is released into the bloodstream during the blood ingestion process. The host immune system produces ROS (Reactive Oxygen Species) as a primary defence mechanism to remove the invading filarial worms. However, well-defined antioxidant enzymes of the nematodes scavenge the host-produced ROS to escape from oxidative stress. The enzyme peroxiredoxin 6 (Prx6) belongs to the peroxiredoxin family, catalyses hydrogen peroxide (H2O2) into water (H2O). In order to find the inhibitors that inhibit the activity of peroxiredoxin 6 of W. bancrofti. We performed the homology modelling to predict the WbPrx6 three-dimensional structure using the Schrödinger-Prime and the dynamic stability of the modelled WbPrx6 was analyzed by carrying out the molecular dynamic (MD) simulation for the time scale of 200ns. Further, the structure-based virtual screening shortlisted the hit molecules from the ChemBridge database based on the glide score. The potential lead molecules (ID: 10239274, 11112883, 79879205, 58160895, and 42133744) that have better binding and satisfied the ADMET properties were selected for further complex simulation and DFT calculations. The identified compounds interact with the N-terminal region of the thioredoxin domain, which plays a key role in reducing phospholipase A2 activity. Interestingly, upon binding the lead molecule, the fluctuation of the loop region that connects α-IV with the ß-VI plays a vital role in affecting the geometry of the active site, which in turn affects the activity WbPrx6. The outcomes of the present computational studies could help in future drug development and designing of the effective candidate to control Lymphatic filariasis.

Molecular evolution, binding site interpretation and functional divergence of aspartate semialdehyde dehydrogenase.

Aspartate Semialdehyde Dehydrogenase (ASDH) is an important enzyme essential for the viability of pathogenic microorganisms. ASDH is mainly involved in amino acid and cell wall biosynthesis of microorganisms, hence it is considered to be a promising target for drug design. This enzyme depicts similar mechanistic function in all microorganisms; although, the kinetic efficiency of an enzyme differs according to their active site residual composition. Therefore, understanding the residual variation and kinetic efficiency of the enzyme would pave new insights in structure-based drug discovery and a novel drug molecule against ASDH. Here, ASDH from Wolbachia endosymbiont of Brugia malayi is used as a prime enzyme to execute evolutionary studies. The phylogenetic analysis was opted to classify 400 sequences of ASDH enzymes based on their structure and electrostatic surfaces. Analysis resulted in 37 monophyletic clades of diverse pathogenic and non-pathogenic organisms. The representative structures of 37 ASDHs from different clades were further deciphered to structural homologues. These enzymes exhibited presence of more positively charged surfaces than negatively charged surfaces in the active site pocket which restrains evolutionary significance. Docking studies of NADP+ with 37 enzymes reveals that site-specific residual variation in the active site pocket modulates the binding affinity (ranges of -13 to -9 kcal/mol). Type-I and Type-II divergence studies show, no significant functional divergence among ASDH, but residual changes were found among the enzyme that modulates the biochemical characteristics and catalytic efficiency. The present study not only explores residual alteration and catalytic variability, it also aids in the design of species-specific inhibitors.Communicated by Ramaswamy H. Sarma.

Computational identification of potential lead molecules targeting rho receptor of Neisseria gonorrhoeae.

Gonorrhea, one of the sexually transmitted disease caused by a gram negative diplococcus bacteria Neisseria gonorrhoeae. Rho protein is indispensable for bacterial viability due to its versatile functions in physiology apart from RNA dependent transcription termination. Based on conserved function and wider role in several cellular processes, inhibitors specifically targeting Rho proteins are largely in use these days to treat various bacterial infections. In this study, three dimensional structure of Rho protein was modeled using the template protein from E. coli and further the optimized model was simulated for 100 ns to understand the structural stability and compactness. Owing to the therapeutic potential of Rho, traditional structure-based virtual screening was applied to identify potential inhibitors for the selected target. Based on empirical glide scoring functions two potent lead molecules (ChemBridge_6121956 and ChemBridge_5232688) were selected from ChemBridge database. The pharmacokinetic properties of these lead molecules are within the permissible range. DFT descriptor revealed that the lead molecules are more reactive, which also supports the molecular docking studies. The stability of Rho and Rho-inhibitor complexes was studied using molecular dynamics simulation. Parameters include binding free energy calculation, RMSD, RMSF and hydrogen bond analysis depicts the stability of Rho and Rho-inhibitors throughout the simulation. Altogether, the identified lead molecules require further optimization towards the design and development of new antibiotics against N. gonorrhoeae.Communicated by Ramaswamy H. Sarma.

Molecular dynamics simulation of Toxin-Antitoxin (TA) system in Acinetobacter baumannii to explore the novel mechanism for inhibition of cell wall biosynthesis: Zeta Toxin as an effective therapeutic target.

The majority of bacteria and archaea contains Toxin-Antitoxin system (TA) that codes for the stable Toxin and unstable Antitoxin components forming a complex. The Antitoxin inhibits the catalytic activities of the Toxin. In general, the Antitoxin will be degraded by the proteases leading to the Toxin activation that subsequently targets essential cellular processes, including transcription, translation, replication, cell division, and cell wall biosynthesis. The Zeta Toxin-Epsilon Antitoxin system in ESKAPE pathogen stabilizes the resistance plasmid and promotes pathogenicity. The known TA system in Acinetobacter baumannii are known to be involved in the replication and translation, however, the mechanism of Zeta Toxin-Epsilon Antitoxin in cell wall biosynthesis remains unknown. In the present study, molecular docking and molecular dynamic (MD) simulations were employed to demonstrate whether Zeta Toxin can impair cell wall synthesis in A. baumannii. Further, the degradation mechanism of Antitoxin in the presence and absence of adenosine triphosphate (ATP) molecules are explained through MD simulation. The result reveals that the cleavage of Antitoxin could be possible with the presence of ATP by displaying its response from 20 ns, whereas the Zeta Toxin/Epsilon was unstable after 90 ns. The obtained results demonstrate that Zeta Toxin is "temporarily favorable" for ATP to undergo phosphorylation at UNAG kinase through the substrate tunneling process. The study further evidenced that phosphorylated UNAG prevents the binding of MurA, the enzyme that catalyzes the initial step of bacterial peptidoglycan biosynthesis. Therefore, the present study explores the binding mechanism of Zeta Toxin/Epsilon Antitoxin, which could be beneficial for preventing cell wall biosynthesis as well as for unveiling the alternative treatment options to antibiotics.

Modelling studies reveal the importance of the C-terminal inter motif loop of NSP1 as a promising target site for drug discovery and screening of potential phytochemicals to combat SARS-CoV-2.

COVID-19 pandemic causative SARS-CoV-2 coronavirus is still rapid in progression and transmission even after a year. Understanding the viral transmission and impeding the replication process within human cells are considered as the vital point to control and overcome COVID-19 infection. Non-structural Protein 1, one among the proteins initially produced upon viral entry into human cells, instantly binds with the human ribosome and inhibit the host translation process by preventing the mRNA attachment. However, the formation of NSP1 bound Ribosome complex does not affect the viral replication process. NSP1 plays an indispensable role in modulating the host gene expression and completely steals the host cellular machinery. The full-length structure of NSP1 is essential for the activity in the host cell and importantly the loop connecting N and C-terminal domains are reported to play a role in ribosome binding. Due to the unavailability of the experimentally determined full-length structure of NSP1, we have modelled the complete structure using comparative modelling and the stability and conformational behaviour of the modelled structure was evaluated through molecular dynamics simulation. Interestingly, the present study reveals the significance of the inter motif loop to serves as a potential binding site for drug discovery experiments. Further, we have screened the phytochemicals from medicinal plant sources since they were used for several hundred years that minimizes the traditional drug development time. Among the 5638 phytochemicals screened against the functionally associated binding site of NSP1, the best five phytochemicals shown high docking score of -9.63 to -8.75 kcal/mol were further evaluated through molecular dynamics simulations to understand the binding affinity and stability of the complex. Prime MM-GBSA analysis gave the relative binding free energies for the top five compounds (dihydromyricetin, 10-demethylcephaeline, dihydroquercetin, pseudolycorine and tricetin) in the range of -45.17 kcal/mol to -37.23 kcal/mol, indicating its binding efficacy in the predicted binding site of NSP1. The density functional theory calculations were performed for the selected five phytochemicals to determine the complex stability and chemical reactivity. Thus, the identified phytochemicals could further be used as effective anti-viral agents to overcome COVID-19 and as well as several other viral infections.