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.

Exploring novel and potent molecules for disrupting DEPTOR-mTOR interaction through structure-steered screening, extra-exactitude molecular docking, prime binding free energy estimation and voguish molecular dynamics.

Dep domain containing mTOR interacting protein (DEPTOR) has critical implications in the development and progression of human malignancies. Increased expression of DEPTOR promotes the growth of tumor cells by inhibiting the mTORC1, which alleviates the negative feedback inhibition by mTORC1 downstream target S6Ks on PI3K/AKT pathway thereby promotes cell survival and prevents apoptosis. This clearly suggests that targetting DEPTOR-mTOR interactions through small molecules may prove as an effective strategy for circumventing distinct cancers. In this study, we employed a top-down approach for finding three novel molecules which may prove effective in disrupting Deptor-mTOR interaction. Following DEPTOR modelling and validation we performed grid-directed structure-based screening by specifying the residues of DEPTOR known to interact with mTOR. A library of 10,000 protein-protein disrupting molecules was screened against the defined region of DEPTOR. From the screened molecules, 30 molecules with highest binding affinity were chosen for molecular docking. Thirty (30) extra-precision molecular docking experiments and 30 molecular mechanics generalized born surface area (MMGBSA) assays were performed. Following this top 10 molecules in terms of binding affinity were selected and the interaction profile of their corresponding docked files was generated. The top three molecules were finally selected after taking all the three parameters including docking score, binding energy value and interaction profile into consideration. For atomistic insights regarding DEPTOR-topmost hit interactions, molecular dynamics was performed for 100 ns. This molecule after further evaluation may prove as promising candidate for anticancer therapy.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.

Delineating binding potential, stability of Sulforaphane-N-acetyl-cysteine in the active site of histone deacetylase 2 and testing its cytotoxicity against distinct cancer lines through stringent molecular dynamics, DFT and cell-based assays.

Histone deacetylase 2 (HDAC2), an isozyme of Class I HDACs has potent imputations in actuating neurodegenerative signaling. Currently, there are sizeable therapeutic disquiets with the use of synthetic histone deacetylase inhibitors in disease management. This strongly suggests the unfulfilled medical necessity of plant substitutes for therapeutic intervention. Sulforaphane-N-acetyl-cysteine (SFN-N-acetylcysteine or SFN-NAC), a sulforaphane metabolite has shown significantly worthier activity against HDACs under in vitro conditions. However, the atomistic studies of SFN-NAC against HDAC2 are currently lacking. Thus, the present study employed a hybrid strategy including extra-precision (XP) grid-based flexible molecular docking, molecular mechanics generalized born surface area (MM-GBSA), e-Pharmacophores method, and molecular dynamics simulation for exploring the binding strengh, mode of interaction, e-Pharmacophoric features, and stability of SFN-NAC towards HDAC2. Further, the globally acknowledged density functional theory (DFT) study was performed on SFN-NAC and entinostat individually in complex state with HDAC2. Apart from this, these inhibitors were tested against three distinct cancer cell models and one transformed cell line for cytotoxic activity. Moreover, double mutant of HDAC2 was generated and the binding orientation and interaction of SFN-NAC was scrutinized in this state. On the whole, this study unbosomed and explained the comparatively higher binding affinity of entinostat for HDAC2 and its wide spectrum cytotoxicity than SFN-NAC.

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.

Evolutionary significance and functional characterization of streptomycin adenylyltransferase from Serratia marcescens.

Complete functional annotations of proteins are essential to understand the role and mechanisms in pathogenesis. Aminoglycoside nucleotidyltransferases are the subclasses of aminoglycosides modifying enzymes conferring resistance to organisms. Insight into the structural and functional understanding of nucleotidyltransferase family protein provides vital information to combat pathogenesis. Phylogenetic analysis is employed to identify the evolutionary significance and common motif's present among the homologs of nucleotidyltransferase family protein. Structure, sequence based approaches and molecular docking were implemented to predict the exact function of the protein. Wide distribution of the nucleotidyltransferase family protein in gram-positive and gram-negative organisms are evidenced from phylogenetic analysis. Five common motifs were present in all the homolog's of nucleotidyltransferase family protein. Sequence-structure based functional annotations predicts that the targeted protein function as ATP-Mg dependent streptomycin adenylyltransferase. Structural comparisons and docking studies correlate well with the identified function. The complete function of nucleotidyltransferase family protein was identified as Streptomycin adenylyltransferase and it could be targeted as a potential therapeutic target to overcome antibiotic resistance.Communicated by Ramaswamy H. SarmaAbbreviationsAACaminoglycoside acetyltransferasesAMEaminoglycoside modifying enzymeANTaminoglycoside nucleotidyltransferasesAPHaminoglycoside phosphotransferasesATPadenosine triphosphateCASTpcomputer atlas and surface topography of proteinsDUFdomains of unknown functionGlidegrid-based ligand docking with energeticHMMhidden Markov modelMASTmotif alignment and search toolMEGAmolecular evolutionary genetics analysisMEMEmultiple Em for motif elicitationMSAmultiple sequence alignmentNMPnucleoside monophosphateNTPnucleoside triphosphateNTnucleotidyltransferaseOPLSoptimized potential for liquid simulationXPextra precision.

Identification of anti-filarial leads against aspartate semialdehyde dehydrogenase of Wolbachia endosymbiont of Brugia malayi: combined molecular docking and molecular dynamics approaches.

Lymphatic filariasis is a debilitating vector borne parasitic disease that infects human lymphatic system by nematode Brugia malayi. Currently available anti-filarial drugs are effective only on the larval stages of parasite. So far, no effective drugs are available for humans to treat filarial infections. In this regard, aspartate semialdehyde dehydrogenase (ASDase) in lysine biosynthetic pathway from Wolbachia endosymbiont Brugia malayi represents an attractive therapeutic target for the development of novel anti-filarial agents. In this present study, molecular modeling combined with molecular dynamics simulations and structure-based virtual screening were performed to identify potent lead molecules against ASDase. Based on Glide score, toxicity profile, binding affinity and mode of interactions with the ASDase, five potent lead molecules were selected. The molecular docking and dynamics results revealed that the amino acid residues Arg103, Asn133, Cys134, Gln161, Ser164, Lys218, Arg239, His246, and Asn321 plays a crucial role in effective binding of Top leads into the active site of ASDase. The stability of the ASDase-lead complexes was confirmed by running the 30 ns molecular dynamics simulations. The pharmacokinetic properties of the identified lead molecules are in the acceptable range. Furthermore, density functional theory and binding free energy calculations were performed to rank the lead molecules. Thus, the identified lead molecules can be used for the development of anti-filarial agents to combat the pathogenecity of Brugia malayi.

Discovery of potent inhibitors targeting Vibrio harveyi LuxR through shape and e-pharmacophore based virtual screening and its biological evaluation.

Quorum sensing is widely recognized as an efficient mechanism in the regulation and production of several virulence factors, biofilm formation and stress responses. For this reason, quorum sensing circuit is emerging as a novel drug target for the development of anti-infective. Recently, cinnamaldehyde derivatives have been found to interfere with master quorum sensing transcriptional regulator and thereby decreasing the DNA binding ability of LuxR. However, the exact mode of cinnamaldehyde binding with LuxR and receptor interaction still remains inconclusive. In the current study, combined method of molecular docking and molecular dynamics simulations were performed to investigate the binding mode, dynamic and energy aspects of cinnamaldehyde derivatives into the binding site of LuxR. Based on the experimental and computational evidences, LuxR-3,4-dichloro-cinnamaldehyde complex was chosen for the development of e-pharmacophore model. Further, shape and e-pharmacophore based virtual screening were performed against ChemBridge database to find potent and suitable ligands for LuxR. By comparing the results of shape and e-pharmacophore based virtual screening; best 9 hit molecules were selected for further studies including ADMET prediction, molecular dynamics simulations and Prime MM-GBSA calculations. From the 9 hit molecules, the top most compound 3-(2,4-dichlorophenyl)-1-(1H-pyrrol-2-yl)-2-propen-1-one (ChemBridge-7364106) was selected for in vitro assays using Vibrio harveyi. The result revealed that ChemBridge-7364106 significantly reduced the bioluminescence production in a dose dependent manner. In addition, ChemBridge-7364106 showed a significant inhibition in biofilm formation and motility in V. harveyi. The results from the study suggest that ChemBridge-7364106 could serve as an anti-quorum sensing molecule for V. harveyi.

Molecular Docking, Molecular Dynamics Simulations, Computational Screening to Design Quorum Sensing Inhibitors Targeting LuxP of Vibrio harveyi and Its Biological Evaluation.

Quorum sensing (QS) plays an important role in the biofilm formation, production of virulence factors and stress responses in Vibrio harveyi. Therefore, interrupting QS is a possible approach to modulate bacterial behavior. In the present study, three docking protocols, such as Rigid Receptor Docking (RRD), Induced Fit Docking (IFD), and Quantum Polarized Ligand Docking (QPLD) were used to elucidate the binding mode of boronic acid derivatives into the binding pocket of LuxP protein in V. harveyi. Among the three docking protocols, IFD accurately predicted the correct binding mode of the studied inhibitors. Molecular dynamics (MD) simulations of the protein-ligand complexes indicates that the inter-molecular hydrogen bonds formed between the protein and ligand complex remains stable during the simulation time. Pharmacophore and shape-based virtual screening were performed to find selective and potent compounds from ChemBridge database. Five hit compounds were selected and subjected to IFD and MD simulations to validate the binding mode. In addition, enrichment calculation was performed to discriminate and separate active compounds from the inactive compounds. Based on the computational studies, the potent Bicyclo [2.2.1] hept-5-ene-2,3-dicarboxylic acid-2,6-dimethylpyridine 1-oxide (ChemBridge_5144368) was selected for in vitro assays. The compound exhibited dose dependent inhibition in bioluminescence and also inhibits biofilm formation in V. harveyi to the level of 64.25 %. The result from the study suggests that ChemBridge_5144368 could serve as an anti-quorum sensing molecule for V. harveyi.

Binding mode exploration of LuxR-thiazolidinedione analogues, e-pharmacophore-based virtual screening in the designing of LuxR inhibitors and its biological evaluation.

Master quorum sensing (QS) regulator LuxR of Vibrio harveyi is a unique member of the TetR protein superfamily. Recent studies have demonstrated the contribution of thiazolidinedione analogues in blocking QS by decreasing the DNA-binding ability of LuxR. However, the precise mechanism of thiazolidinedione analogues binding to LuxR is still unclear. In the present study, molecular docking combined with molecular dynamics (MD) simulations was performed to understand the mechanism of ligand binding to the protein. The binding pattern of thiazolidinedione analogues showed strong hydrogen bonding interactions with the amine group (NH) of polar amino acid residue Asn133 and carbonyl (C=O) interaction with negatively charged amino acid residue Gln137 in the binding site of LuxR. The stability of the protein-ligand complexes was confirmed by running 50 ns of MD simulations. Further, the four-featured pharmacophore hypothesis (AHHD) consists of one acceptor (A), two hydrophobic regions (HH) and one donor (D) group was used to screen compounds from ChemBridge database. The identified hit molecules were shown to have excellent pharmacokinetic properties under the acceptable range. Based on the computational studies, ChemBridge_5343641 was selected for in vitro assays. The 1-(4-chlorophenoxy)-3-[(4,6-dimethyl-2-pyrimidinyl)thio]-2-propanol (ChemBridge_5343641) showed significant reduction in bioluminescence in a dose-dependent manner. In addition, ChemBridge_5343641 inhibits biofilm formation and motility in V. harveyi. The result from the study suggests that ChemBridge_5343641 could serve as an anti-QS molecule.

Pharmacophore modeling and structure-based virtual screening to identify potent inhibitors targeting LuxP of Vibrio harveyi.

The main aim of the study is to identify molecules that can disrupt quorum sensing (QS) system of Vibrio harveyi and therefore perhaps the production of toxins. Recently, a novel class of dioxazaborocane derivatives has been found to block AI-2 QS by targeting LuxPQ, but the mechanism of protein inhibition is still unclear. In order to investigate the possible binding modes, all the derivatives were docked into the binding site of LuxP using induced fit docking (IFD). The computed binding affinity is in good agreement with the experimental data. Resultant protein-ligand complexes were simulated using Desmond module and the result revealed better binding of ligands in the binding site of LuxP. Both pharmacophore- and structure-based virtual screening was performed to identify novel hits against LuxP. A filtering protocol, including lipinski filters, number of rotatable bonds and three levels of docking precisions were used for the selection of hits with specific properties. The virtual screening results were then combined and analyzed, which retrieved six hits with significant Glide score, binding affinity toward LuxP. The pharmacokinetic properties of the retrieved hits are in the acceptable range. Enrichment calculation was performed to validate the final hits, to discriminate the active compounds from the inactive compounds. The identified hits could serve as a base for further drug development against LuxP of Vibrio harveyi.

Protective effect of Mollugo nudicaulis Lam. on acute liver injury induced by perchloroethylene in experimental rats.

OBJECTIVE: To evaluate the protective effect of ethanol extract of Mollugo nudicaulis (M. nudicaulis) against perchloroethylene-induced hepatotoxicity. METHODS: The hepatoprotective activity of the ethanol extract of M. nudicaulis (200 mg/kg body wt) was studied in percholoroethylene (1 000 mg/kg body wt) induced hepatotoxicity in Wistar albino rats. The serum levels of AST, ALT, ALP, bilirubin and the liver content of SOD, CAT, GPx, GST, GSH, vitamin C were assessed to evaluate the hepatoprotective and antioxidant activities of the extract. The activity of the extract was compared with silymarin, a standard reference drug. In addition, serum urea, uric acid and creatinine levels were measured to evaluate the kidney function. The histopathological examination of the liver tissues was observed to support the biochemical parameters. RESULTS: The results revealed that the extract significantly (P<0.05) restored the serum levels of AST, ALT, ALP, bilirubin and significantly (P<0.05) increased the antioxidant enzymes SOD, CAT, GPx, GST, GSH, vitamin C in perchloroethylene-induced rats to its normalcy. The biochemical observations were supported by the histopathological studies of the liver tissues. CONCLUSIONS: The results led to the conclusion that M. nudicaulis possess hepatoprotective and antioxidant activities against perchloroethylene-induced hepatotoxicity in rats.