In silico docking and molecular dynamics simulation of 3-dehydroquinate synthase (DHQS) from <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis>

The shikimate pathway is as an attractive target because it is present in bacteria, algae, fungi, and plants but does not occur in mammals. In Mycobacterium tuberculosis (MTB), the shikimate pathway is integral to the biosynthesis of naphthoquinones, menaquinones, and mycobactin. In these study, novel inhibitors of 3-dehydroquinate synthase (DHQS), an enzyme that catalyzes the second step of the shikimate pathway in MTB, were determined. 12,165 compounds were selected from two public databases through virtual screening and molecular docking analysis using PyRx 8.0 and Autodock 4.2, respectively. A total of 18 compounds with the best binding energies (?13.23 to ?8.22 kcal/mol) were then selected and screened for absorption, distribution, metabolism, excretion, and toxicity (ADMET) analysis, and nine of those compounds were found to satisfy all of the ADME and toxicity criteria. Among those nine, the three compounds—ZINC633887?(binding energy =??10.29 kcal/mol), ZINC08983432?(?9.34 kcal/mol), and PubChem73393?(?8.61 kcal/mol)—with the best binding energies were further selected for molecular dynamics (MD) simulation analysis. The results of the 50-ns MD simulations showed that the two compounds ZINC633887 and PubChem73393 formed stable complexes with DHQS and that the structures of those two ligands remained largely unchanged at the ligand-binding site during the simulations. These two compounds identified through docking and MD simulation are potential candidates for the treatment of TB, and should undergo validation in vivo and in vitro.     

Development of gallic acid formazans as novel enoyl acyl carrier protein reductase inhibitors for the treatment of tuberculosis

The enoyl acyl carrier protein reductase (InhA) of Mycobacterium tuberculosis (MTB) is an attractive target for developing novel antitubercular agents. A series of gallic acid formazans, were computationally designed and docked into the active site of InhA to understand their binding mode and potential to inhibit InhA. Nine compounds from the designed series were identified as potential InhA inhibitors, on the basis of good Glide score. These compounds were synthesized in the laboratory and evaluated for in vitro antitubercular activity against drug-sensitive and multi-drug resistant strains of MTB. Out of nine compounds, three compounds exhibited the most promising MIC of <2 μM against the sensitive strain of MTB, H37Rv. The compounds were evaluated against five resistant strains of MTB. Most of the compounds exhibited activity superior to the standard, linezolid, against all these resistant strains. The mechanism of action of these compounds was concluded to be InhA inhibition, through InhA enzyme inhibition study. Insignificant cytotoxicity of these compounds was observed on RAW 264.7 cell line. Inactivity of all these compounds against gram positive and gram negative bacteria indicated their specificity against MTB. The compounds were further analyzed for ADME properties and showed potential as good oral drug candidates. The results clearly identified some novel, selective and specific InhA inhibitors against sensitive and resistant strains of MTB.     

Structure-based screening and molecular dynamics simulations offer novel natural compounds as potential inhibitors of Mycobacterium tuberculosis isocitrate lyase

Mycobacterium tuberculosis is the etiological agent of tuberculosis in humans and is responsible for more than two million deaths annually. M. tuberculosis isocitrate lyase (MtbICL) catalyzes the first step in the glyoxylate cycle, plays a pivotal role in the persistence of M. tuberculosis, which acts as a potential target for an anti-tubercular drug. To identify the potential anti-tuberculosis compound, we conducted a structure-based virtual screening of natural compounds from the ZINC database (n = 1,67,748) against the MtbICL structure. The ligands were docked against MtbICL in three sequential docking modes that resulted in 340 ligands having better docking score. These compounds were evaluated for Lipinski and ADMET prediction, and 27 compounds were found to fit well with re-docking studies. After refinement by molecular docking and drug-likeness analyses, three potential inhibitors (ZINC1306071, ZINC2111081, and ZINC2134917) were identified. These three ligands and the reference compounds were further subjected to molecular dynamics simulation and binding energy analyses to compare the dynamic structure of protein after ligand binding and the stability of the MtbICL and bound complexes. The binding free energy analyses were calculated to validate and capture the intermolecular interactions. The results suggested that the three compounds had a negative binding energy with ?96.462, ?143.549, and ?122.526 kJ mol^?1 for compounds with IDs ZINC1306071, ZINC2111081, and ZINC2134917, respectively. These lead compounds displayed substantial pharmacological and structural properties to be drug candidates. We concluded that ZINC2111081 has a great potential to inhibit MtbICL and would add to the drug discovery process against tuberculosis.     

Structural and energetic understanding of novel natural inhibitors of Mycobacterium tuberculosis malate synthase

Persistent infection by Mycobacterium tuberculosis requires the glyoxylate shunt. This is a bypass to the tricarboxylic acid cycle in which isocitrate lyase (ICL) and malate synthase (MS) catalyze the net incorporation of carbon during mycobacterial growth on acetate or fatty acids as the primary carbon source. To identify a potential antitubercular compound, we performed a structure-based screening of natural compounds from the ZINC database (n = 1 67 740) against the M tuberculosis MS (MtbMS) structure. The ligands were screened against MtbMS, and 354 ligands were found to have better docking score. These compounds were assessed for Lipinski and absorption, distribution, metabolism, excretion, and toxicity prediction where 15 compounds were found to fit well for redocking studies. After refinement by molecular docking and drug-likeness analysis, four potential inhibitors (ZINC1483899, ZINC1754310, ZINC2269664, and ZINC15729522) were identified. These four ligands with phenyl-diketo acid were further subjected to molecular dynamics simulation to compare the dynamics and stability of the protein structure after ligand binding. The binding energy analysis was calculated to determine the intermolecular interactions. Our results suggested that the four compounds had a binding free energy of −201.96, −242.02, −187.03, and −169.02 kJ·mol⁻¹, for compounds with IDs ZINC1483899, ZINC1754310, ZINC2269664, and ZINC15729522, respectively. We concluded that two compounds (ZINC1483899 and ZINC1754310) displayed considerable structural and pharmacological properties and could be probable drug candidates to fight against M tuberculosis parasites.     

Computational finding of potential inhibitor for Cytochrome P450 Mono-oxygenases Enzyme of Mycobacterium tuberculosis

Cytochrome P450 mono-oxygenases (2UUQ) enzyme from Mycobacterium tuberculosis catalyzes oxidation of organic compounds such as lipids and steroidal hormones therefore remain as potential drug target. Currently available first line anti-tuberculosis drugs have been caused several side effects in the body as well as resistance development by mycobacterium against these drugs, necessitates the considerable need for finding new drugs. Therefore, we propose a structure based computational method to find a new potential inhibitor for cytochrome P450 mono-oxygenases enzyme. Compounds from several ligand databases were docked against the functional sites of 2UUQ (A) through the standard GEMDOCK v2.0 and AUTODOCK4.0 molecular docking tools. Commercially available chemical compound ZINC00004165 (5-[3-(2-nitroimidazol-1-yl) propyl] phenanthridine) has produced top rank with lowest interaction energy of -113.2 (via GEMDOCK) and lowest docking energy of -9.80 kcal/mol (via AUTODOCK) as compared to first line anti TB compounds. Z score and normal distribution analysis verified that the ZINC00004165 compound has more affinity towards 2UUQ in comparison to large number of random population of compounds. ZINC00004165 is also in agreement with the drug likeness properties of Lipinski rule of five without any violation. Therefore, our finding concludes that the commercial compound ZINC00004165 can act as a potential inhibitor against cytochrome P450 mono-oxygenases enzyme of Mycobacterium tuberculosis.     

Repurposing of FDA-approved drugs to target MurB and MurE enzymes in Mycobacterium tuberculosis

Abstract

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb) is one amongst the top 10 causes of death worldwide. The growing rise in antibiotic resistance compounded with slow and expensive drug discovery has further aggravated the situation. ‘Drug repurposing’ is a promising approach where known drugs are examined for a new indication. In the present study, we have attempted to identify drugs that could target MurB and MurE enzymes involved in the muramic acid synthesis pathway (Mur Pathway) in Mtb. FDA-approved drugs from two repositories i.e. Drug Bank (1932 drugs) and e-LEA3D (1852 drugs) were screened against these proteins. Several criteria were applied to study the protein-drug interactions and the consensus drugs were further studied by molecular dynamics (MD) simulation. Our study found Sulfadoxine (–7.3?kcal/mol) and Pyrimethamine (–7.8?kcal/mol) to show stable interaction with MurB while Lifitegrast (–10.5?kcal/mol) and Sildenafil (–9.1?kcal/mol) showed most reliable interaction with MurE. Furthermore, binding free energy (ΔG_bind), RMSD and RMSF data and the number of hydrogen bonds corroborated the stability of interactions and hence these drugs for repurposing should be explored further.

Communicated by Ramaswamy H. Sarma     

Ursolic acid and carvacrol may be potential inhibitors of dormancy protein small heat shock protein16.3 of Mycobacterium tuberculosis

Small heat shock protein16.3 (sHSP16.3) is a crucial protein for survival of Mycobacterium tuberculosis (MTB) in its host. Besides, this protein acts as a molecular chaperone during stress and is indispensable for MTB’s growth, virulence and cell-wall thickening. sHSP16.3 is also a promising candidate for vaccine, serodiagnosis and drug design as well. In the present study, we have targeted sHSP16.3 with two phytochemicals, namely ursolic acid and carvacrol using in silico approach. Molecular docking analysis showed that both phytochemicals (ursolic acid and carvacrol) have docked with sHSP16.3 and shown tendency to inhibit the function of this vital protein of MTB. In addition, both compounds have exhibited strong compatibility with sHSP16.3 during whole 60 ns duration of molecular dynamics simulation. Further, the molecular mechanic/generalized Born/Poisson–Boltzmann surface area (MM/G/P/BSA) free energies were calculated which showed that both phytocompounds have stable and favourable binding energies causing strong binding with binding site of sHSP16.3. Taking together, the data of present study suggest that both phytocompounds may be potential inhibitor of sHSP16.3 of MTB and a best alternative to standard anti-tuberculosis drugs.     

Metabolic pathway analysis and molecular docking analysis for identification of putative drug targets in Toxoplasma gondii: novel approach

Toxoplasma gondii is an obligate intracellular apicomplexan parasite that can infect a wide range of warm-blooded animals including humans. In humans and other intermediate hosts, toxoplasma develops into chronic infection that cannot be eliminated by host's immune response or by currently used drugs. In most cases, chronic infections are largely asymptomatic unless the host becomes immune compromised. Thus, toxoplasma is a global health problem and the situation has become more precarious due to the advent of HIV infections and poor toleration of drugs used to treat toxoplasma infection, having severe side effects and also resistance have been developed to the current generation of drugs. The emergence of these drug resistant varieties of T. gondii has led to a search for novel drug targets. We have performed a comparative analysis of metabolic pathways of the host Homo sapiens and the pathogen T. gondii. The enzymes in the unique pathways of T. gondii, which do not show similarity to any protein from the host, represent attractive potential drug targets. We have listed out 11 such potential drug targets which are playing some important work in more than one pathway. Out of these, one important target is Glutamate dehydrogenase enzyme; it plays crucial part in oxidation reduction, metabolic process and amino acid metabolic process. As this is also present in the targets of tropical diseases of TDR (Tropical disease related Drug) target database and no PDB and MODBASE 3D structural model is available, homology models for Glutamate dehydrogenase enzyme were generated using MODELLER9v6. The model was further explored for the molecular dynamics simulation study with GROMACS, virtual screening and docking studies with suitable inhibitors against the NCI diversity subset molecules from ZINC database, by using AutoDock-Vina. The best ten docking solutions were selected (ZINC01690699, ZINC17465979, ZINC17465983, ZINC18141294_03, ZINC05462670, ZINC01572309, ZINC18055497_01, ZINC18141294, ZINC05462674 and ZINC13152284_01). Further the Complexes were analyzed through LIGPLOT. On the basis of Complex scoring and binding ability it is deciphered that these NCI diversity set II compounds, specifically ZINC01690699 (as it has minimum energy score and one of the highest number of interactions with the active site residue), could be promising inhibitors for T. gondii using Glutamate dehydrogenase as Drug target.     

Molecular Interaction of Novel Compound 2-Methylheptyl Isonicotinate Produced by Streptomyces sp. 201 with Dihydrodipicolinate Synthase (DHDPS) Enzyme of Mycobacterium tuberculosis for its Antibacterial Activity

Antibiotic resistance is a growing problem in multi-drug-resistant tuberculosis which is caused by Mycobacterium tuberculosis (MTB). Hence there is an urgent need for designing or developing a novel or potent anti-tubercular agent. The Lysine/DAP biosynthetic pathway is a promising target because of its role in cell wall and amino acid biosynthesis. In our study we performed a molecular docking analysis of a novel antibacterial isolated from Streptomyces sp. 201 at three different binding site of dihydrodipicolinate synthase (DHDPS) enzyme of MTB. The molecular docking studies suggest that the novel molecule shows favourable interaction at the three different binding sites as compared to five experimentally known inhibitors of DHDPS.     

In vitro anti-TB properties,in silico target validation,molecular docking and dynamics studies of substituted 1,2,4-oxadiazole analogues against Mycobacterium tuberculosis

The alarming increase in multi- and extensively drug-resistant (MDR and XDR) strains of Mycobacterium tuberculosis (MTB) has triggered the scientific community to search for novel, effective, and safer therapeutics. To this end, a series of 3,5-disubstituted-1,2,4-oxadiazole derivatives (3a–3i) were tested against H37Rv, MDR and XDR strains of MTB. Of which, compound 3a with para-trifluorophenyl substituted oxadiazole showed excellent activity against the susceptible H37Rv and MDR-MTB strain with a MIC values of 8 and 16 µg/ml, respectively.To understand the mechanism of action of these compounds (3a–3i) and identify their putative drug target, molecular docking and dynamics studies were employed against a panel of 20 mycobacterial enzymes reported to be essential for mycobacterial growth and survival. These computational studies revealed polyketide synthase (Pks13) enzyme as the putative target. Moreover, in silico ADMET predictions showed satisfactory properties for these compounds, collectively, making them, particularly compound 3a, promising leads worthy of further optimisation.     

Identification of potential inhibitors of Fasciola gigantica thioredoxin1: computational screening,molecular dynamics simulation,and binding free energy studies

Fasciola gigantica is the causative organism of fascioliasis and is responsible for major economic losses in livestock production globally. F. gigantica thioredoxin1 (FgTrx1) is an important redox-active enzyme involved in maintaining the redox homeostasis in the cell. To identify a potential anti-fasciolid compound, we conducted a structure-based virtual screening of natural compounds from the ZINC database (n = 1,67,740) against the FgTrx1 structure. The ligands were docked against FgTrx1 and 309 ligands were found to have better docking score. These compounds were evaluated for Lipinski and ADMET prediction, and 30 compounds were found to fit well for re-docking studies. After refinement by molecular docking and drug-likeness analysis, three potential inhibitors (ZINC15970091, ZINC9312362, and ZINC9312661) were identified. These three ligands were further subjected to molecular dynamics simulation (MDS) to compare the dynamics and stability of the protein structure after binding of the ligands. The binding free energy analyses were calculated to determine the intermolecular interactions. The results suggested that the two compounds had a binding free energy of –82.237, and –109.52 kJ.mol^?1 for compounds with IDs ZINC9312362 and ZINC9312661, respectively. These predicted compounds displayed considerable pharmacological and structural properties to be drug candidates. We concluded that these two compounds could be potential drug candidates to fight against F. gigantica parasites.     

Drug targeted virtual screening and molecular dynamics of LipU protein of Mycobacterium tuberculosis and Mycobacterium leprae

The lipolytic protein LipU was conserved in mycobacterium sp. including M. tuberculosis (MTB LipU) and M. leprae (MLP LipU). The MTB LipU was identified in extracellular fraction and was reported to be essential for the survival of mycobacterium. Therefore to address the problem of drug resistance in pathogen, LipU was selected as a drug target and the viability of finding out some FDA approved drugs as LipU inhibitors in both the cases was explored. Three-dimensional (3D) model structures of MTB LipU and MLP LipU were generated and stabilized through molecular dynamics (MD). FDA approved drugs were screened against these proteins. The result showed that the top-scoring compounds for MTB LipU were Diosmin, Acarbose and Ouabain with the Glide XP score of ?12.8, ?11.9 and ?11.7 kcal/mol, respectively, whereas for MLP LipU protein, Digoxin (?9.2 kcal/mol), Indinavir (?8.2 kcal/mol) and Travoprost (?8.2 kcal/mol) showed highest affinity. These drugs remained bound in the active site pocket of MTB LipU and MLP LipU structure and interaction grew stronger after dynamics. RMSD, RMSF and Rg were found to be persistent throughout the simulation period. Hydrogen bonds along with large number of hydrophobic interactions stabilized the complex structures. Binding free energies obtained through Prime/MM-GBSA were found in the significant range from ?63.85 kcal/mol to ?34.57 kcal/mol for MTB LipU and ?71.33 kcal/mol to ?23.91 kcal/mol for MLP LipU. The report suggested high probability of these drugs to demolish the LipU activity and could be probable drug candidates to combat TB and leprosy disease.     

Identification of novel natural inhibitors of Opisthorchis felineus cytochrome P450 using structure-based screening and molecular dynamic simulation

Opisthorchis felineus is the etiological agent of opisthorchiasis in humans. O. felineus cytochrome P450 (OfCYP450) is an important enzyme in the parasite xenobiotic metabolism. To identify the potential anti-opisthorchid compound, we conducted a structure-based virtual screening of natural compounds from the ZINC database (n = 1,65,869) against the OfCYP450. The ligands were screened against OfCYP450 in four sequential docking modes that resulted in 361 ligands having better docking score. These compounds were evaluated for Lipinski and ADMET prediction, and 10 compounds were found to fit well with re-docking studies. After refinement by docking and drug-likeness analyses, four potential inhibitors (ZINC2358298, ZINC8790946, ZINC70707116, and ZINC85878789) were identified. These ligands with reference compounds (itraconazole and fluconazole) were further subjected to molecular dynamics simulation (MDS) and binding energy analyses to compare the dynamic structure of protein after ligand binding and the stability of the OfCYP450 and bound complexes. The binding energy analyses were also calculated. The results suggested that the compounds had a negative binding energy with ?259.41, ?110.09, ?188.25, ?163.30, ?202.10, and ?158.79 kJ mol^?1 for itraconazole, fluconazole, and compounds with IDs ZINC2358298, ZINC8790946, ZINC70707116, and ZINC85878789, respectively. These lead compounds displayed significant pharmacological and structural properties to be drug candidates. On the basis of MDS results and binding energy analyses, we concluded that ZINC8790946, ZINC70707116, and ZINC85878789 have excellent potential to inhibit OfCYP450.     

Virtual screening, identification and in vitro testing of novel inhibitors of O-acetyl-L-serine sulfhydrylase of Entamoeba histolytica

The explosive epidemicity of amoebiasis caused by the facultative gastrointestinal protozoan parasite Entamoeba histolytica is a major public health problem in developing countries. Multidrug resistance and side effects of various available antiamoebic drugs necessitate the design of novel antiamobeic agents. The cysteine biosynthetic pathway is the critical target for drug design due to its significance in the growth, survival and other cellular activities of E. histolytica. Here, we have screened 0.15 million natural compounds from the ZINC database against the active site of the EhOASS enzyme (PDB ID. 3BM5, 2PQM), whose structure we previously determined to 2.4 Å and 1.86 Å resolution. For this purpose, the incremental construction algorithm of GLIDE and the genetic algorithm of GOLD were used. We analyzed docking results for top ranking compounds using a consensus scoring function of X-Score to calculate the binding affinity and using ligplot to measure protein-ligand interactions. Fifteen compounds that possess good inhibitory activity against EhOASS active site were identified that may act as potential high affinity inhibitors. In vitro screening of a few commercially available compounds established their biological activity. The first ranked compound ZINC08931589 had a binding affinity of ∼8.05 µM and inhibited about 73% activity at 0.1 mM concentration, indicating good correlation between in silico prediction and in vitro inhibition studies. This compound is thus a good starting point for further development of strong inhibitors.     

A novel class of antimicrobial drugs selectively targets a Mycobacterium tuberculosis PE-PGRS protein

The continued spread of drug-resistant tuberculosis is one of the most pressing and complex challenges facing tuberculosis management worldwide. Therefore, developing a new class of drugs is necessary and urgently needed to cope with the increasing threat of drug-resistant tuberculosis. This study aims to discover a potential new class of tuberculosis drug candidates different from existing tuberculosis drugs. By screening a library of compounds, methyl (S)-1-((3-alkoxy-6,7-dimethoxyphenanthren-9-yl)methyl)-5-oxopyrrolidine-2-carboxylate (PP) derivatives with antitubercular activity were discovered. MIC ranges for PP1S, PP2S, and PP3S against clinically isolated drug-resistant Mycobacterium tuberculosis strains were 0.78 to 3.13, 0.19 to 1.56, and 0.78 to 6.25 μg/ml, respectively. PPs demonstrated antitubercular activities in macrophage and tuberculosis mouse models, showing no detectable toxicity in all assays tested. PPs specifically inhibited M. tuberculosis without significantly changing the intestinal microbiome in mice. Mutants selected in vitro suggest that the drug targets the PE-PGRS57, which has been found only in the genomes of the M. tuberculosis complex, highlighting the specificity and safety potency of this compound. As PPs show an excellent safety profile and highly selective toxicity specific to M. tuberculosis, PPs are considered a promising new candidate for the treatment of drug-resistant tuberculosis while maintaining microbiome homeostasis.

This study identifies a new class of PP derivatives as Mycobacterium tuberculosis-targeting antimicrobials with microbiome-safe properties that do not disrupt normal flora, making them a promising candidate for the treatment of drug-resistant tuberculosis.     

Trypanosoma cruzi: Activity of heterocyclic cationic molecules in vitro

Chagas disease remains a serious public health problem in several Latin American countries. New chemotherapy is urgently needed since current drugs are limited in efficacy and exhibit undesirable side effects. Aromatic diamidines and analogs are well known anti-parasitic agents and in this study, we have evaluated the in vitro trypanocidal effect of several different heterocyclic cationic compounds, including diamidines (DB1195, DB1196 and DB1345), a monoamidine (DB824), an arylimidamide (DB613A) and a guanylhydrazone (DB1080) against amastigotes and bloodstream trypomastigotes of Trypanosoma cruzi, the etiological agent of Chagas disease. Our present findings showed that all compounds exerted, at low-micromolar doses, a trypanocidal effect upon both intracellular parasites and bloodstream trypomastigotes of T. cruzi. The activity of DB1195, DB1345, DB824 and DB1080 against bloodstream forms was reduced when these compounds were assayed in the presence of mouse blood possibly due to their association with plasma constituents and/or due to metabolic instability of the compounds. However, trypanocidal effects of DB613A and DB1196 were not affected by plasma constituents, suggesting their potential application in the prophylaxis of banked blood. In addition, potency and selectivity of DB613A, towards intracellular parasites, corroborate previous results that demonstrated the highly promising activity of arylimidamides against this parasite, which justify further studies in experimental models of T. cruzi infection.     

New 1,4-di-N-oxide-quinoxaline-2-ylmethylene isonicotinic acid hydrazide derivatives as anti-Mycobacterium tuberculosis agents

The increase in the prevalence of drug-resistant tuberculosis cases demonstrates the need of discovering new and promising compounds with antimycobacterial activity. As a continuation of our research and with the aim of identifying new antitubercular drugs candidates, a new series of quinoxaline 1,4-di-N-oxide derivatives containing isoniazid was synthesized and evaluated for in vitro anti-tuberculosis activity against Mycobacterium tuberculosis H37Rv strain. Moreover, various drug-like properties of new compounds were predicted. Taking into account the biological results and the promising drug-likeness profile of these compounds, make them valid leads for further experimental research.     

Design,synthesis, and bioevaluation of a novel class of (E)-4-oxo-crotonamide derivatives as potent antituberculosis agents

A series of novel (E)-4-oxo-2-crotonamide derivatives were designed and synthesized to find potent antituberculosis agents. All the target compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis H₃₇Rv(MTB). Results reveal that 4-phenyl moiety at part A and short methyl group at part C were found to be favorable. Most of the derivatives displayed promising activity against MTB with MIC ranging from 0.125 to 4?µg/mL. Especially, compound IIIa16 was found to have the best activity with MIC of 0.125?μg/mL against MTB and with MIC in the range of 0.05–0.48?µg/mL against drug-resistant clinical MTB isolates.