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Acetyl Coenzyme A Carboxylase (AccD6) is a homodimeric protein which is involved in the carboxylation of acetyl coenzyme A to produce malonyl coenzyme A, which plays an important role in the biosyn...

NAD synthetase (NadE) catalyzes the last step in NAD biosynthesis, transforming deamido-NAD+ into NAD+ by a two-step reaction with co-substrates ATP and amide donor ammonia. In this study, we report the crystal structure of Staphylococcus aureus NAD synthetase enzyme (saNadE) at 2.3 A resolution. We used this structure to perform molecular dynamics simulations of apo-enzyme, enzyme-substrate (NadE with ATP and NaAD) and enzyme-intermediate complexes (NadE with NaAD-AMP) to investigate key binding interactions and explore the conformational transitions and flexibility of the binding pocket. Our results show large shift of N-terminal region in substrate bound form which is important for ATP binding. Substrates drive the correlated movement of loop regions surrounding it as well as some regions distal to the active site and stabilize them at complex state. Principal component analysis of atomic projections distinguish feasible trajectories to delineate distinct motions in enzyme-substrate to enzyme-intermediate states. Our results suggest mixed binding involving dominant induced fit and conformational selection. MD simulation extracted ensembles of NadE could potentially be utilized for in silico screening and structure based design of more effective Methicillin Resistant Staphylococcus aureus (MRSA) inhibitors.

Trypanothione reductase of Leishmania donovani is a flavin adenine dinucleotide containing homodimeric protein essential for parasite survival. The flavoenzyme utilizes nicotinamide adenine dinucleotide phosphate in the reaction to convert oxidized trypanothione to reduced trypanothione which is further used up by tryparedoxin/tryparedoxin peroxidase system to neutralize the reactive oxygen species generated by the macrophages. Some of the drugs previously reported against the disease include sodium stibogluconate, miltefosine and amphotericin B. However, due to the resistance and toxicity problem associated with these molecules, there is an urgent need to develop new drugs against L. donovani. Trypanothione reductase of L. donovani is one such essential target whose inhibition could lead to a decline in parasite growth. In this work, we have performed a computational studies using Maybridge library of chemical compounds to identify potential inhibitors of Trypanothione reductase of L. donovani. Structure-based virtual screening method in combination with molecular docking was employed to identify and prioritize 30 compounds which were further subjected to molecular dynamics simulation. Ten compounds which showed stable ligand root-mean-square deviation plot, c-alpha backbone and root-mean-square fluctuation were considered for trypanothione reductase inhibition assay and subsequent inhibition studies of parasite growth. Enzyme inhibition assay resulted in shortlisting of four compounds that were found to inhibit Trypanothione reductase of L. donovani. Subsequently, the anti-leishmanial screening highlighted one compound as the potential anti-leishmanial agent, with IC50 value of 15.2 µM, that can be further optimised with medicinal chemistry efforts to improve its activity. Communicated by Ramaswamy H. Sarma

Over the last few decades, anticancer peptides (ACPs) have turned into potential warheads against cancer. Apart from small molecules and monoclonal antibodies, ACPs have been proven to be effective against cancer cells. ACPs are small cationic peptides that selectively bind to the negatively charged cancer cell membrane and kill them by various mechanisms. In the present study, we prepared a random scrambled library of 1200 peptides from the 100 known ACPs and virtually screened them for their anticancer properties. From in silico-predicted ACPs, 27 peptides were prioritized based on their support vector machine (SVM) score. Based on the SVM score and properties such as hydrophobicity, size, overall net charge, secondary structure, and synthetic feasibility, finally, four peptides were synthesized and screened for their biological activities. Cancer cell membrane-deforming potential of two most active peptides, peptide1 and peptide2 was assessed with molecular dynamics simulation. We found that peptide1 remains adsorbed to the membrane surface, while peptide2 has membrane penetration capability. The present study will be helpful in the computational design of ACPs and understanding their interaction with the cancerous cell's membrane.

The malaria parasite carries two organelles, the apicoplast and mitochondrion, whose DNA genomes must be maintained for optimal function and parasite survival under genotoxic stress. DNA repair mechanism(s) operative within these organelles were explored by mining the Plasmodium falciparum nuclear genome for sequences encoding proteins of major DNA repair pathways with predicted targeting to either organelle. Of the panel of enzymes identified for base excision repair (BER), we characterized the apurinic/apyrimidinic (AP) endonuclease PfApn1-an EndoIV whose homolog is not known in humans. PfApn1 targeted to the mitochondrion and functioned as an AP endonuclease requiring both Zn2+ and Mn2+ ions for maximal activity. Mutation of the critical third metal-binding site residue H542 resulted in the loss of Mn2+ (but not Zn2+ ) binding indicating that Mn2+ bound PfApn1 at this site; this was further supported by molecular dynamic simulation. CD spectra analysis further showed requirement of both metal ions for the attainment of PfApn1 β-strand-rich optimal conformation. PfApn1 also functioned as a 3'-phosphatase that would enable removal of 3'-blocks for DNA polymerase activity during BER. Interestingly, unlike Escherichia coli and yeast EndoIV homologs, PfApn1 lacked 3'-5' exonuclease activity and also did not cleave damaged bases by nucleotide incision repair (NIR). Uncoupling of endonuclease/phosphatase and exonuclease/NIR in PfApn1 suggests that amino acid residues distinct from those critical for endonuclease function are required for exonuclease activity and NIR. Characterization of a critical mitochondrion-targeted AP endonuclease provides evidence for a functional BER pathway in the parasite organelle.

Cervical cancer is a leading cause of cancer-related deaths among women in developing countries. Therefore, development of new chemotherapeutic agents is required. Unlike normal cells, cancer cells contain elevated copper levels which play an integral role in angiogenesis. Thus, targeting copper via copper-specific chelators in cancer cells can serve as effective anticancer strategy. In this work, a copper chelator pregnenolone acetate nucleus-based tetrazole derivative (ligand-L) was synthesized and characterized by elemental analysis, ESI-MS, 1H NMR and 13C NMR. DNA binding ability of ligand-L was studied using UV–Vis and fluorescence spectroscopy. Fluorescence spectroscopy studies reveal that quenching constant of ligand- l -DNA and ligand-L-Cu(II) were found to be 7.4 × 103 M−1 and 8.8 × 103 M−1, respectively. In vitro toxicity of ligand-L was studied on human cervical cancer C33A cancer cells. Results showed that ligand-L exhibit significant cytotoxic activity against cervical cancer C33A cells with IC50 value 5.0 ± 1.8 µM. Further, it was found that ligand-L cytotoxicity is due to redox cycling of copper to generate ROS which leads to DNA damage and apoptosis. In conclusion, this is the report where we synthesized pregnenolone acetate-based tetrazole derivative against C33A cells that targets cellular copper to induce pro-oxidant death in cancer cells. These findings will provide significant insights into the development of new chemical molecules with better copper chelating and pro-oxidant properties against cancer cells.

The synthesis of 5-formyl-6-aryl-6H-indolo[3,2,1-de][1,5] naphthyridine-2-carboxylates by reaction between 1-formyl-9H-β-carbolines and cinnamaldehydes in the presence of pyrrolidine in water with microwave irradiation is described. Pharmacophoric modification of the formyl group offered several new fused β-carboline derivatives, which were investigated for their κ-opioid receptor (KOR) agonistic activity. Two compounds 4 a and 4 c produced appreciable agonist activity on KOR with EC50 values of 46±19 and 134±9 nM, respectively. Moreover, compound-induced KOR signaling studies suggested both compounds to be extremely G-protein-biased agonists. The analgesic effect of 4 a was validated by the increase in tail flick latency in mice in a time-dependent manner, which was completely blocked by the KOR-selective antagonist norBNI. Moreover, unlike U50488, an unbiased full KOR agonist, 4 a did not induce sedation. The docking of 4 a with the human KOR was studied to rationalize the result.

Background Defects in DNA repair pathway can lead to double-strand breaks leading to genomic instability. Earlier we have shown that S.pombe Drp1, a Rint1/Tip1 family protein is required for the recovery from DNA damage. Methods Various truncations of Drp1 protein were constructed and their role in DNA damage response and interaction with Rad50 protein has been studied by co-immunoprecipitation and pull-down assays. Results The structural and functional analysis of Drp1 protein revealed that the N-terminus region of Drp1 is indispensable for the survival. The C-terminus truncation mutants, drp1C1Δ and drp1C2Δ exhibit temperature sensitive phenotype and are hypersensitive against DNA damaging agents with elevated level of Rad52-YFP foci at non-permissive temperature indicating the impairment for DNA damage repair pathway. The essential N-terminus region of Drp1 interacts with the C-terminus region of Rad50 and might be involved in influencing the MRN/X function. Small-angle X-ray (SAXS) analysis revealed three-domain like shapes in Drp1 protein while the C-terminus region of Rad50 exhibit unusual bulges. Computational docking studies revealed the amino acid residues at the C-terminus region of Rad50 that are involved in the interaction with the residues present at the N-terminal region of Drp1 indicating the importance of the N-terminal region of Drp1 protein. Conclusions We have identified the region of Drp1 and Rad50 proteins that are involved in the interaction and their role in the DNA damage response pathway has been analyzed. General significance The functional and structural aspects of fission yeast Drp1 protein and its interaction with Rad50 have been elucidated.

Unique intrinsic properties of peptides like low toxicity, high biological activity, and specificity make them attractive therapeutic agents. PDZ-binding peptide inhibitors have been demonstrated for curing of Alzheimer, Parkinson, Dementia, and other central nervous system ailments. In this article, we report the successful use of an integrated computational protocol to analyze the structural basis of how peptides bind to the shallow groove of the third PDZ domain (PDZ-3) from the postsynaptic density (PSD-95) protein. This protocol employs careful and precise computational techniques for design of new strategy for predicting novel and potent peptides against PDZ protein. We attempted to generate a pharmacophore model using crystal structure of peptide inhibitor bound to the PDZ-3. A highly specific and sensitive generated pharmacophore model was used for screening virtual database generated using different combination of amino acid substitutions as well as decoy peptide database for its sensitivity and specificity. Identified hit peptides were further analyzed by docking studies, and their stability analyzed using solvated molecular dynamics. Quantum Mechanics/Molecular Mechanics (QM/MM) interaction energy and GMX-PBSA scoring schemes were used for ranking of stable peptides. Computational approach applied here generated encouraging results for identifying peptides against PDZ interaction model. The workflow can be further exercised as a virtual screening technique for reducing the search space for candidate target peptides against PDZ domains.

A concise, one pot and regioselective access to (E)-distyrylbenzenes (DSBs) from arylhalide and secondary phenylenediethanol, a stable precursor for in situ generation of divinylbenzene (DVB) to avoid its polymerization, is described for construction of double C C bond formation via tandem double-dehydrative-double-Heck (D-D-D-H) reaction using Palladium and ionic liquid [hmim]Br as a cooperative catalyst. It is noteworthy that this pot-economy approach also provides direct synthesis of hydroxylated distyrylbenzenes without requirement of protection-deprotection strategy. Importantly, the synthesized DSBs are tested for their protective activity against β amyloid reduction, acetylcholine esterase inhibition, lipid lowering and reactive oxygen species (ROS) reduction properties in transgenic Caenorhabditis elegans model wherein 1,3-bis((E)-4-(trifluoromethyl)styryl)benzene (5c) is found to be active across all above factors thus presenting lead molecule within multi-target-directed ligands (MTDLs) approach. Molecular docking studies were also performed to understand the interactions of potent DSBs with receptors.

In an endeavor to search for affordable and safer therapeutics against debilitating visceral leishmaniasis, we examined antileishmanial potential of ammonium trichloro [1,2-ethanediolato-O,O']-tellurate (AS101); a tellurium based non toxic immunomodulator. AS101 showed significant in vitro efficacy against both Leishmania donovani promastigotes and amastigotes at sub-micromolar concentrations. AS101 could also completely eliminate organ parasite load from L. donovani infected Balb/c mice along with significant efficacy against infected hamsters (˃93% inhibition). Analyzing mechanistic details revealed that the double edged AS101 could directly induce apoptosis in promastigotes along with indirectly activating host by reversing T-cell anergy to protective Th1 mode, increased ROS generation and anti-leishmanial IgG production. AS101 could inhibit IL-10/STAT3 pathway in L. donovani infected macrophages via blocking α4β7 integrin dependent PI3K/Akt signaling and activate host MAPKs and NF-κB for Th1 response. In silico docking and biochemical assays revealed AS101's affinity to form thiol bond with cysteine residues of trypanothione reductase in Leishmania promastigotes leading to its inactivation and inducing ROS-mediated apoptosis of the parasite via increased Ca2+ level, loss of ATP and mitochondrial membrane potential along with metacaspase activation. Our findings provide the first evidence for the mechanism of action of AS101 with excellent safety profile and suggest its promising therapeutic potential against experimental visceral leishmaniasis.

Dipeptidyl peptidase 4 (DPP4) is a well-known target for the antidiabetic drugs. However, currently available DPP4 inhibitor screening assays are costly and labor-intensive. It is important to create a robust in silico method to predict the activity of DPP4 inhibitor for the new lead finding. Here, we introduce an R-based Web application SVMDLF (SVM-based DPP4 Lead Finder) to predict the inhibitor of DPP4, based on support vector machine (SVM) model, predictions of which are confirmed by in vitro biological evaluation. The best model generated by MACCS structure fingerprint gave the Matthews correlation coefficient of 0.87 for the test set and 0.883 for the external test set. We screened Maybridge database consisting approximately 53,000 compounds. For further bioactivity assay, six compounds were shortlisted, and of six hits, three compounds showed significant DPP4 inhibitory activities with IC50 values ranging from 8.01 to 10.73 μm. This application is an OpenCPU server app which is a novel single-page R-based Web application for the DPP4 inhibitor prediction. The SVMDLF is freely available and open to all users at http://svmdlf.net/ocpu/library/dlfsvm/www/ and http://www.cdri.res.in/svmdlf/.

Nucleoside diphosphate kinases (NDKs) are ubiquitous enzymes that catalyze the transfer of the γ-phosphate moiety from an NTP donor to an NDP acceptor, crucial for maintaining the cellular level of nucleoside triphosphates (NTPs). The inability of trypanosomatids to synthesize purines de novo and their dependence on the salvage pathway makes NDK an attractive target to develop drugs for the diseases they cause. Here we report the discovery of novel inhibitors for Leishmania NDK based on the structural and functional characterization of purified recombinant NDK from Leishmania amazonensis. Recombinant LaNDK possesses auto-phosphorylation, phosphotransferase and kinase activities with Histidine 117 playing an essential role. LaNDK crystals were grown by hanging drop vapour diffusion method in a solution containing 18% PEG-MME 500, 100 mM Bis-Tris propane pH 6.0 and 50 mM MgCl2. It belongs to the hexagonal space group P6322 with unit cell parameters a = b = 115.18, c = 62.18 A and α = β = 90°, γ = 120°. The structure solved by molecular replacement methods was refined to crystallographic R-factor and Rfree values of 22.54 and 26.52%, respectively. Molecular docking and dynamics simulation-based virtual screening identified putative binding compounds. Protein inhibition studies of selected hits identified five inhibitors effective at micromolar concentrations. One of the compounds showed ~45% inhibition of Leishmania promastigotes proliferation. Analysis of inhibitor-NDK complexes reveals the mode of their binding, facilitating design of new compounds for optimization of activities as drugs against leishmaniasis.

Glutamine synthetase is an enzyme that catalyzes the condensation of glutamate and ammonia to form glutamine in the presence of adenosine triphosphate. The wealth of structure-related information about glutamine synthetase across the species is endorsing it as emerging potential drug target. Owing to its well-characterized role in metabolism of Mycobacterium, various high throughput screening studies have been aimed at the identification of inhibitors against MtGS. The present work is focussed on comparative sequence and structural studies of glutamine synthetase and its evaluation as drug target against the infectious diseases. We have done molecular modeling studies of glutamine synthetase of Leishmania and Plasmodium. The structure models and molecular dynamics simulations studies shed light on to the binding modes of substrates viz. adenosine diphosphate, glutamate, ammonia, and metal ions. The comparative studies of MtGS, HsGS, LmGS, and PvGS helped in better understanding of prospects of structure-based inhibitor design. The results suggest that amino acid-binding site is highly conserved, whereas nucleotide-binding site possess subtle variations and thus offers opportunity for specific inhibitor design. Therefore, present study suggests that broad spectrum glutamine synthetase inhibition is feasible and it is potential drug target against infectious diseases.

This study describes the isolation and purification of a phytocystatin from seeds of Brassica juncea (Indian mustard; cultivar RoAgro 5444), which is an important oilseed crop both agriculturally and economically. The protein was purified by gel filtration chromatography with 24.3% yield and 204-fold purification, and visualised by 2D gel electrophoresis. The 18.1 kDa mustard cystatin was highly specific for cysteine proteinases. The plant cystatin inhibited cathepsin B, confirming its role in conferring pest resistance. The inhibitor was highly stable over a pH range of 3–10 and retained significant inhibitory potential up to 70 °C. The stoichiometry of its interaction with papain, determined by isothermal calorimetry, suggests a 1:1 complex. Secondary structural elements calculated by far-UV circular dichroism (CD) spectroscopy show an 18.8% α-helical and 21% β-sheet structure. The protein was a non-competitive inhibitor of thiol proteinases. The Stokes radius and frictional co-efficient were used to describe the shape and size of the protein. Homology modelling and docking studies proposed a prototype illustrating the Brassica phytocystatin mediated papain inhibition. Molecular dynamics (MD) study revealed the excellent stability of the papain–phytocystatin complex during a simulation for 100 ns. Detailed results identify the mustard cystatin as an important member of the phytocystatin family.

Brugia malayi Glucose 6-phosphate dehydrogenase apoenzyme (BmG6PD) was expressed and purified by affinity chromatography to study the differences in kinetic properties of enzyme and the effect of the cofactor NADP(+) binding on enzyme stability. The presence of cofactor NADP(+) influenced the tertiary structure of enzyme due to significant differences in the tryptophan microenvironment. However, NADP(+) binding have no effect on secondary structure of the enzyme. Quenching with acrylamide indicated that two or more tryptophan residues became accessible upon cofactor binding. Unfolding and cross linking study of BmG6PD showed that NADP(+) stabilized the protein in presence of high concentration of urea/GdmCl. A homology model of BmG6PD constructed using human G6PD (PDB id: 2BH9) as a template indicated 34% α-helix, 19% β-sheet and 47% random coil conformations in the predicted model of the enzyme. In the predicted model binding of NADP(+) to BmG6PD was less tight with the structural sites (-10.96 kJ/mol binding score) as compared with the coenzyme site (-15.47 kJ/mol binding score).

In this study, a series of coumarin–indole hybrids have been synthesized and evaluated for their lipid lowering activity. Preliminary biological screening of the synthesized compounds was undertaken in an in vitro model of the HMG-CoA reductase enzyme, and the activity was confirmed in Triton WR-1339 induced hyperlipidemic rats. Among the hybrids, compound 26 was found to be the best as it significantly reduced the serum and hepatic lipid profiles in an HFD-fed hyperlipidemic rat model. The mechanism of action seems to be associated with the regulation of HMG-CoA reductase activity in the liver, which is in good agreement with binding mode studies. Compound 26 exhibited favorable pharmacokinetic behavior for its oral administration, which underscores the potential of this template as a new class of hypolipidemic agents.

In the quest for novel scaffolds for the management of androgen sensitive prostatic disorders like prostate cancer and benign prostatic hyperplasia, a series of twenty-six aryl/heteroaryl piperazine derivatives have been described. Three compounds, 8a, 8c and 9a, exhibited good activity profiles against an androgen sensitive prostate cancer cell line (LNCaP) with EC50 values of 9.8, 7.6 and 11.2 μM, respectively. These compounds caused a decrease in luciferase activity and a decline in PSA and Ca2+ levels, which are indicative of their anti-androgenic and α1A-adrenergic receptor blocking activities, respectively. Compound 9a reduced the prostate weight of rats (47%) and in pharmacokinetic analysis at 10 mg kg−1 it demonstrated an MRT of ∼14 h post dose, exhibiting high levels in prostate. Compound 9a docked in a similar orientation to hydroxyflutamide on an androgen receptor and showed strong π–π interactions. These findings reveal that compound 9a is a promising candidate for management of prostatic disorders with anti-androgenic and α1A-blocking activities.

A series of new dihydropyrimidinone–semicarbazone hybrids were successfully synthesised by integrating regioselective multicomponent reaction with the pharmacophore hybridization approach. All the synthesised compounds were evaluated for their hLig1 inhibition potency and most of them were found to be good to moderately active. Out of the tested derivatives, compound 6f showed selective anti-proliferative activity against HepG2 cells in a dose-dependent manner with an IC50 value of 10.07 ± 1.2. It also reduced cell survival at ≤20 μM concentration. Further, analysis of treated HepG2 cell lysates by western blot assay showed increased γ-H2AX levels and upregulation of p53, leading to apoptosis. In silico docking results explain the binding modes of compound 6f to the DNA-binding domain of hLig1 enzyme thereby preventing its nick sealing activity. In addition, the favourable pharmacokinetic properties suggest that this new class of hLig1 inhibitors could be promising leads for further drug development.

Recent advances indicate a possible role of phytochemicals as modulatory factors in signaling pathways. We have previously demonstrated PHLPP2-mediated suppression of Nrf2 responses during oxidant attack. The present study was designed to explore Nrf2-potentiating mechanism of morin, a flavonol, via its possible role in intervening PHLPP2-regulated Akt/GSK3β/Fyn kinase axis. Efficacy of morin was evaluated against oxidative stress-mediated damage to primary hepatocytes by tert-butyl hydroperoxide (tBHP) and acetaminophen. The anti-cytotoxic effects of morin were found to be a consequence of fortification of Nrf2-regulated antioxidant defenses since morin failed to sustain activities of redox enzyme in Nrf2 silenced hepatocytes. Morin promoted Nrf2 stability and its nuclear retention by possibly modulating PHLPP2 activity which subdues cellular Nrf2 responses by activating Fyn kinase. Pull-down assay using morin-conjugated beads indicated the binding affinity of morin towards PHLPP2. Molecular docking also revealed the propensity of morin to occupy the active site of PHLPP2 enzyme. Thus, dietary phytochemical morin was observed to counteract oxidant-induced hepatocellular damage by promoting Nrf2-regulated transcriptional induction. The findings support the novel role of morin in potentiating Nrf2 responses by limiting PHLPP2 and hence Fyn kinase activation. Therefore, morin may be exploited in developing novel therapeutic strategy aimed at enhancing Nrf2 responses., Graphical abstract fx1, Highlights • Cytoprotection against tBHP-evoked oxidative stress by morin is Nrf2-regulated. • Morin prevents Nrf2-destabilization via PHLPP2-Akt-GSK3β-Fyn kinase pathway. • In silico docking studies confirmed PHLPP2 activity inhibition by morin. • Morin mitigates APAP induced cytotoxicity by suppressing PHLPP2 pathway.

Enoyl-acyl carrier protein reductase (InhA) of type II fatty acid synthase system is involved in the synthesis of mycolic acids which is a major component of the bacterial cell wall. Since they are the key enzymes playing a very significant role in the FASII pathway of the bacterium. In this study, we have developed a workflow for identification of InhA inhibitors by utilizing in silico virtual screening approaches based on various machine learning algorithms followed by pharmacophore based virtual screening. The hits screened from the models were further subjected to molecular docking. Further, based on the XP docking score best twenty compounds were subjected to molecular dynamics study. Finally, nine compounds were shortlisted on the basis of best stable ligand RMSD, c-alpha RMSD, and RMSF plot for biological evaluation studies. Experimental validation of the shortlisted compounds identified one compound JFD01724 having potent inhibitory activity and was able to inhibit the growth of mycobacterium tuberculosis. Further medicinal chemistry efforts may help to improve the inhibitory potency of the identified compound.

Twenty-four novel benzofuran-pyran derivatives were synthesized and evaluated for their anti-osteoporotic activity in primary cultures of rat calvarial osteoblasts in vitro. Among all the compounds screened for the alkaline phosphatase activity, three compounds 4e, 4j and 4k showed potent activity at picomolar concentrations in osteoblast differentiating stimulation. Additionally, these compounds were found effective in mineralization, assessed by alizarin red-S staining assay. Compounds were again validated through a series of other in vitro experiments. Moreover, molecular dynamics simulations demonstrated that both benzofuran and pyran moieties are requisite to fit into the active site of BMP-2 receptor, a key target of the osteogenic agents. The obtained results strongly convey that compound 4e is a potential bone anabolic agent among synthesized series, which can be further explored as a drug lead for treating osteoporosis.

Vaccination is devised/formulated to stimulate specific and prolonged immune responses for long-term protection against infection or disease. A vaccine component, namely adjuvant, enhances antigen recognition by the host immune system and thereby stimulates its cellular and adaptive responses. Especially synthetic Toll-like receptor (TLR) agonists having self-assembling properties are considered as good candidates for adjuvant development. Here, a human TLR4-derived 20-residue peptide (TR-433), present in the dimerization interface of the TLR4–myeloid differentiation protein-2 (MD2) complex, displayed self-assembly and adopted a nanostructure. Both in vitro studies and in vivo experiments in mice indicated that TR-433 is nontoxic. TR-433 induced pro-inflammatory responses in THP-1 monocytes and HEK293T cells that were transiently transfected with TLR4/CD14/MD2 and also in BALB/c mice. In light of the self-assembly and pro-inflammatory properties of TR-433, we immunized with a mixture of TR-433 and either ovalbumin or filarial antigen trehalose-6-phosphate phosphatase (TPP). A significant amount of IgG titers was produced, suggesting adjuvanting capability of TR-433 that was comparable with that of Freund's complete adjuvant (FCA) and appreciably higher than that of alum. We found that TR-433 preferentially activates type 1 helper T cell (T(h)1) response rather than type 2 helper T cell (T(h)2) response. To our knowledge, this is the first report on the identification of a short TLR4-derived peptide that possesses both self-assembling and pro-inflammatory properties and has significant efficacy as an adjuvant, capable of activating cellular responses in mice. These results indicate that TR-433 possesses significant potential for development as a new adjuvant in therapeutic application.

Combating tuberculosis requires new therapeutic strategies that not only target the actively dividing bacilli but also the dormant bacilli during persistent infection. Isocitrate lyase (ICL) is a key enzyme of the glyoxylate shunt, crucial for the survival of bacteria in macrophages and mice. MtbICL is considered as one of the potential and attractive drug targets against persistent infection. We report the inhibition of MtbICL by quercetin with IC 50 of 3.57 μM. In addition, quercetin strongly inhibited the growth of Mtb H37Rv utilizing acetate, rather than glucose as the sole carbon source, suggesting the inhibition of glyoxylate shunt. Quercetin binds at the N-terminus of MtbICL ( K d – 6.68 μM).

Isocitrate lyase (ICL), is a key enzyme of the glyoxylate shunt crucial for the survival of Mycobacterium tuberculosis (Mtb) in macrophages during persistent infection. MtbICL catalyses the first step of this carbon anaplerosis cycle and is considered as a potential anti-tubercular drug target. The MtbICL is a tetramer with 222 symmetry, and each subunit of the enzymeis composed of 14 α-helices and 14 β-strands. We studied the conformational flexibility of the enzyme to get a deeper insight into its stability and function. Our studies show that the mutation of His180, close to the MtbICL signature sequence (K193KCGH197) completely abolishes the oligomeric conformation and function of the enzyme. Molecular dynamics studies suggest that the loss of interaction between His180 and Tyr89 most likely alters the orientation of Tyr89 side chain, thereby causing the movement of helices α6, α12, α13 and α14 in the vicinity and affecting the tetrameric assembly. We further show that the oligomerization of MtbICL is primarily mediated by the inter subunit interactions, and strengthened by the helix swapping of α12–α13 between adjacent subunits. Furthermore, the enzyme activity is influenced by the interactions between the residues of lid region (P411NSSTTALTGSTEEGQFH428) and the loop region (T391KHQREV397). Mutation of glutamates of the lid region to non homologous residues (E423A or E424A) or basic residues (E423K or E424K) inactivates the enzyme, whereas the activity is not much compromised in case of homologous mutations (E423D or E424D).

Prophylactic prevention is considered as the most promising strategy to tackle STI/HIV. Twenty-five dithiocarbamate-thiourea hybrids (14-38) were synthesized as woman controlled topical vaginal microbicides to counter Trichomonas vaginalis and sperm along with RT inhibition potential. The four promising compounds (18, 26, 28 and 33) were tested for safety through cytotoxic assay against human cervical cell line (HeLa) and compatibility with vaginal flora, Lactobacillus. Docking study of most promising vaginal microbicide (33) revealed that it docked in a position and orientation similar to known reverse transcriptase inhibitor Nevirapine. The preliminary in vivo pharmacokinetics of compound 33 was performed in NZ-rabbits to evaluate systemic toxicity in comparison to Nonoxynol-9.

DNA replication and repair are complex processes accomplished by the concerted action of a network of enzymes and proteins. DNA ligases play a crucial role in these processes by catalyzing the nick joining between DNA strands. As compared to normal cells, elevated levels of human DNA ligase I (hLigI) is reported in some cancers. We studied the inhibition of hLigI mediated DNA nick sealing activity followed by the antiproliferative activity of novel indole-chalcone based benzopyran compounds on cancer cells. One molecule called compound 27 showed a notable preference for inhibition of hLigI as compared to other ligases and showed enhanced cytotoxicity against colon cancer (DLD-1) cells as compared to normal cells. Mechanistic studies showed that compound 27 directly interacts with hLigI in a competitive manner and did not interact with the DNA substrate during ligation. This novel and potent hLigI inhibitor showed significant inhibition of both monolayer culture as well as 3D culture of DLD-1 cells that mimic solid tumor. It also affected the migration of DLD-1 cells indicating the potential anti-metastatic activity. This novel hLigI inhibitor could therefore serve as a promising lead for anticancer drug development.

A series of novel benzofuran-dihydropyridine hybrids were designed by molecular hybridization approach and evaluated for bone anabolic activities. Among the screened library, ethyl 4-(7-(sec-butyl)-2-(4-methylbenzoyl)benzofuran-5-yl)-2-methyl-5-oxo-1,4,5,6,7,8-hexahydroquinoline-3-carboxylate (compound 21) significantly enhanced the ALP production and mineralized nodule formation, which are primary requisites in the process of in vitro osteogenesis. Oral administration of compound 21 at 10 mg.kg−1 day−1 for two weeks led to restoration of trabecular bone microarchitecture in drill hole fracture model by significantly increasing BV/TV and Tb.N. Furthermore, histological and molecular studies showed compound 21 triggering the new bone regeneration in a drill hole defect site by increasing BMP expression. Furthermore, molecular modeling studies were performed to gain insight into the binding approach, which revealed that both benzofuran and dihydropyridine moieties are essential to show similar binding interactions to fit into the active site of BMP2 receptor, an important target of the osteogenic agents. Our results suggest that compound 21 stimulates BMP2 synthesis in osteoblast cells that promotes new bone formation (∼40%) at the fracture site which helps in shorten the healing period.

Cathepsin S has been demonstrated to play a crucial role in the remodeling of extracellular matrix proteins such as elastin and collagen, which in turn contribute to the structural integrity of the cardiovascular wall. Atherosclerotic lesions, aneurysm formation, plaque rupture, thrombosis, and calcification are some of the cardiovascular disorders related to cathepsin S. A highly selective inhibitor of human as well as animal cathepsin S, RO5444101, was recently reported to attenuate the progression of atherosclerotic lesions. Here, we attempted to gain insight into the molecular mechanism of action of RO5444101 on cathepsin S by performing molecular docking and molecular dynamics (MD) simulation studies. The results of our studies correlate well with relevant reported experimental data and potentially explain the selectivity of this inhibitor for cathepsin S rather than cathepsin L1/L, cathepsin L2/V, and cathepsin K, which share conserved catalytic sites and have sequence similarities of 49%, 50%, and 55%, respectively, with respect to cathepsin S. In contrast to those closely related cathepsins, 20 ns MD simulation data reveal that the overall interaction of cathepsin S with RO5444101 is more stable and involves more protein-molecule interactions than the interactions of the inhibitor with the other cathepsins. This study therefore considerably improves our understanding of the molecular mechanism responsible for cathepsin S inhibition and facilitates the identification of potential novel selective inhibitors of cathepsin S.

The relict plastid (apicoplast) of the malaria parasite is the site for important biochemical pathways and is essential for parasite survival. The SUF pathway of [Fe-S] cluster assembly in the apicoplast of Plasmodium spp. is of interest due to its absence in the human host suggesting the possibility of anti-malarial intervention through apicoplast [Fe-S] biogenesis. We report biochemical characterization of components of the Plasmodium falciparum apicoplast SUF pathway after the first step of sulfur mobilization. In vitro interaction experiments and in-vivo crosslinking showed that apicoplast-encoded PfSufB and apicoplast-targeted PfSufC and PfSufD formed a complex. The PfSufB-C2-D complex could function as a scaffold to assemble [4Fe-4S] clusters in vitro and activity of the PfSufC ATPase was enhanced by PfSufD. Two carrier proteins- the NifU-like protein PfNfu and the A-type carrier PfSufA are homodimers, the former mediating transfer of [4Fe-4S] from the scaffold to a model [4Fe-4S] target protein with higher efficiency. Conditional knockout of SufS, the enzyme catalysing the first step of sulfur mobilization, by selective excision in the mosquito stages of P. berghei severely impaired development of sporozoites in oocysts establishing essentiality of the SUF machinery in the vector. Our results delineate steps of the complete apicoplast SUF pathway and demonstrate its critical role in the parasite life cycle. This article is protected by copyright. All rights reserved.

In the search for effective multifunctional agents for the treatment of Alzheimer's disease (AD), a series of novel hybrids incorporating benzofuran and chalcone fragments were designed and synthesized. These hybrids were screened by using a transgenic Caenorhabditis elegans model that expresses the human β-amyloid (Aβ) peptide. Among the hybrids investigated, (E)-3-(7-methyl-2-(4-methylbenzoyl)benzofuran-5-yl)-1-phenylprop-2-en-1-one (4 f), (E)-3-(2-benzoyl-7-methylbenzofuran-5-yl)-1-phenylprop-2-en-1-one (4 i), and (E)-3-(2-benzoyl-7-methylbenzofuran-5-yl)-1-(thiophen-2-yl)prop-2-en-1-one (4 m) significantly decreased Aβ aggregation and increased acetylcholine (ACh) levels along with the overall availability of ACh at the synaptic junction. These compounds were also found to decrease acetylcholinesterase (AChE) levels, reduce oxidative stress in the worms, lower lipid content, and to provide protection against chemically induced cholinergic neurodegeneration. Overall, the multifunctional effects of these hybrids qualify them as potential drug leads for further development in AD therapy.

A series of multifunctional directed 3-arylcoumarin-tetracyclic tacrine derivatives was designed and synthesized for the treatment of Parkinson's disease (PD). A number of derivatives (18, 19, 20, 21, and 24) demonstrated significant reduction of aggregation of "human" alpha-synuclein (α-synuclein) protein, expressing on transgenic Caenorhabditis elegans (C. elegans) model NL5901. Moreover, compounds 16, 18, and 24 also exhibited good antioxidant properties and significantly increased the dopamine (DA) content in N2 and NL5901 strains of C. elegans. Interestingly, the protective efficacy of these hybrids seems to be mediated via activation of longevity promoting transcription factor DAF-16. In addition, molecular modeling studies have evidenced the exquisite interaction of most active compounds 18 and 24 with α-synuclein protein. Taken together, the data indicate that the derivatives may be useful leads against aging and age associated PD.

The plastid of the malaria parasite, the apicoplast, is essential for parasite survival. It houses several pathways of bacterial origin that are considered attractive sites for drug intervention. Among these is the sulfur mobilization (SUF) pathway of Fe-S cluster biogenesis. Although the SUF pathway is essential for apicoplast maintenance and parasite survival, there has been limited biochemical investigation of its components and inhibitors of Plasmodium SUFs have not been identified. We report the characterization of two proteins, Plasmodium falciparum SufS ( Pf SufS) and Pf SufE, that mobilize sulfur in the first step of Fe-S cluster assembly and confirm their exclusive localization to the apicoplast. The cysteine desulfurase activity of Pf SufS is greatly enhanced by Pf SufE, and the Pf SufS- Pf SufE complex is detected in vivo . Structural modeling of the complex reveals proximal positioning of conserved cysteine residues of the two proteins that would allow sulfide transfer from the PLP (pyridoxal phosphate) cofactor-bound active site of Pf SufS. Sulfide release from the l -cysteine substrate catalyzed by Pf SufS is inhibited by the PLP inhibitor d -cycloserine, which forms an adduct with Pf SufS-bound PLP. d -Cycloserine is also inimical to parasite growth, with a 50% inhibitory concentration close to that reported for Mycobacterium tuberculosis , against which the drug is in clinical use. Our results establish the function of two proteins that mediate sulfur mobilization, the first step in the apicoplast SUF pathway, and provide a rationale for drug design based on inactivation of the PLP cofactor of Pf SufS.

Thymidylate kinase (TMK) is a potential chemotherapeutic target because it is directly involved in the synthesis of deoxythymidine triphosphate, which is an essential component for DNA synthesis. The gene encoding thymidylate kinase of Brugia malayi was amplified by PCR and expressed in Escherichia coli. The native molecular weight of recombinant B. malayi thymidylate kinase (rBmTMK) was estimated to be ∼52 kDa by gel filtration chromatography, suggesting a homodimeric structure. rBmTMK activity required divalent cation and Mg2+ was found to be the most effective cation. The enzyme was sensitive to pH and temperature, it showed maximum activity at pH 7.4 and 37 °C. The Km values for dTMP and ATP were 17 and 66 μM, respectively. The turnover number kcat was found to be 38.09 s−1, a value indicating the higher catalytic efficiency of the filarial enzyme. The nucleoside analogues 5-bromo-2′-deoxyuridine (5-BrdU), 5-chloro-2′-deoxyuridine (5-CldU) and 3′-azido-3′-deoxythymidine (AZT) showed specific inhibitory effect on the enzyme activity and these effects were in good association with binding interactions and the scoring functions as compared to human TMK. Differences in kinetic properties and structural differences in the substrate binding site of BmTMK model with respect to human TMK can serve as basis for designing specific inhibitors against parasitic enzyme.

The reduced genomes of the apicoplast and mitochondrion of the malaria parasite Plasmodium falciparum are actively translated and antibiotic-mediated translation inhibition is detrimental to parasite survival. In order to understand recycling of organellar ribosomes, a critical step in protein translation, we identified ribosome recycling factors (RRF) encoded by the parasite nuclear genome. Targeting of PfRRF1 and PfRRF2 to the apicoplast and mitochondrion respectively was established by localization of leader sequence-GFP fusions. Unlike any RRF characterized thus far, PfRRF2 formed dimers with disulphide interaction(s) and additionally localized in the cytoplasm, thus suggesting adjunct functions for the factor. PfRRF1 carries a large 108-amino-acid insertion in the functionally critical hinge region between the head and tail domains of the protein, yet complemented Escherichia coli RRF in the LJ14frr(ts) mutant and disassembled surrogate E. coli 70S ribosomes in the presence of apicoplast-targeted EF-G. Recombinant PfRRF2 bound E. coli ribosomes and could split monosomes in the presence of the relevant mitochondrial EF-G but failed to complement the LJ14frr(ts) mutant. Although proteins comprising subunits of P. falciparum organellar ribosomes are predicted to differ from bacterial and mitoribosomal counterparts, our results indicate that the essential interactions required for recycling are conserved in parasite organelles.

The ethanolic extract and fractions of Wrightia t omentosa Roem. & Schult (Apocynaceae) leaves were tested in vivo for their antidyslipidemic activity in high fat diet (HFD) induced dyslipidemic hamsters. Activity guided isolation resulted in identification of antidyslipidemic compounds β-AA and β-AP. Compounds β-AA and β-AP decrease the levels of LDL by 36% and 44%, and increase the HDL-C/TC ratio by 49% and 28%, respectively, at a dose of 10 mg/kg. In addition, the isolated compounds β-AA and β-AP showed significant HMG-CoA-reductase inhibition, which was further established by docking studies.

A series of 35 triazolopyrimidine analogues reported as Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) inhibitors were optimized using quantum mechanics methods, and their binding conformations were studied by docking and 3D quantitative structure–activity relationship studies. Genetic algorithm-based criteria was adopted for selection of training and test sets while maintaining structural diversity of training and test sets, which is also very crucial for model development and validation. Both the comparative molecular field analyses (\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ q_{\text{LOO}}^2 = 0.{841} $$\end{document}, \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ r_{\text{ncv}}^2 = 0.{99} $$\end{document}) and comparative molecular similarity indices analyses (\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ q_{\text{LOO}}^2 = 0.{757} $$\end{document}, \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ r_{\text{ncv}}^2 = 0.{943} $$\end{document}) show excellent correlation and high predictive power. Furthermore, molecular dynamics simulations were performed to explore the binding mode of the two of the most active compounds of the series, 10 and 14. Harmonization in the two simulation results validate the analysis and therefore applicability of docking parameters based on crystallographic conformation of compound 14 bound to receptor molecule. This work provides useful information about the inhibition mechanism of this class of molecules and will assist in the design of more potent inhibitors of PfDHODH.

Current treatment of leishmaniasis is based on chemotherapy, which relies on a handful of drugs with serious limitations, such as high cost, toxicity, and lack of efficacy in endemic regions. Therefore, development of new, effective, and affordable anti-leishmanial drugs is a global health priority. Dipeptidylcarboxypeptidase has been characterized and established as a drug target for antileishmanial drug discovery. We virtually screened a large chemical library of 15 452 compounds against a 3D model of dipeptidylcarboxypeptidase to identify novel inhibitors. The initial virtual screening using a ligand-based pharmacophore model identified 103 compounds. Forty-six compounds were shortlisted based on the docking scores and other scoring functions. Further, these compounds were subjected to biological assay, and four of them belonging to two chemical classes were identified as the lead compounds. Identification of these novel and chemically diverse inhibitors should provide leads to be optimized into candidates to treat these protozoan infections.

Bioassay guided fractionation of the ethanolic extract of Polyalthia longifolia var. pendula, led to the discovery of the clerodane diterpene, 16α-hydroxycleroda-3, 13 (14) Z-dien-15, 16-olide (1), as a new structural class of HMG-CoA reductase inhibitor. Importantly, the in vivo effects of 1 corroborated well with its molecular docking analysis and also with its hamster plasma pharmacokinetics.

The interaction of a well characterized new complex 1 cis,fac-[RuCl(dmso-S)3(L)] LH = 1-(2-hydroxyphenyl)-3-(4-chlorophenyl) propenone with Calf-thymus DNA (CT-DNA) is monitored using UV–vis titration (Kb = 3.8 × 107) and ethidium bromide displacement studies (Ksv = 3.2). The molecular docking of the complex with DNA sequence d(ACCGACGTCGGT)2 reveals that complex is stabilized by additional electrostatic and hydrogen bonding interaction with DNA besides probable displacement of a labile DMSO by the N7 of guanine. The coordination of guanine is further supported by the isolation and characterization of its adduct with the complex 1.gua (gua = guanine). The nuclease property of complex 1 in the absence and presence of different activators and trappers demonstrates that complex efficiently cleaves supercoiled pBR322 plasmid DNA and binds through major groove of the DNA.

Thymidylate synthase (TS) is a crucial enzyme for DNA biosynthesis and many nonclassical lipophilic antifolates targeting this enzyme are quite efficient and encouraging as antitumor drugs. Herein, we report some 3D-QSAR analyses using CoMFA and CoMSIA on quinozoline antifolates in order to have a better understanding of the mechanism of action and structure-activity relationship of these compounds. By applying leave-one-out (LOO) cross-validation study, we obtained cross-validated q2 value of 0.573 for CoMFA and 0.445 for CoMSIA, while the non-cross-validated r2 values for them were found to be 0.935 and 0.893, respectively. The models were graphically interpreted using CoMFA and CoMSIA contour plots. The results obtained from this study could be used for rational design of potent inhibitors against thymidylate synthase.

Transketolase, the most critical enzyme of the non-oxidative branch of the pentose phosphate pathway, has been reported as a novel target in Plasmodium falciparum as it has least homology with the human host. Homology model of P. falciparum transketolase (PfTk) was constructed using the crystal structure of S. cervisiae transketolase as a template, and used for the identification and prioritization of potential compounds targeted against Plasmodium falciparum transketolase. The docking studies with fructose-6-phosphate and thiamine pyrophosphate showed that His31, Asp473, Ser388, Arg361 and His465 formed hydrogen bonds with fructose-6-phosphate while pyrimidine ring of coenzyme interacted with conserved residues of protein viz., Leu121, Glu415, Gly119. The major interacting residues involved in binding of oxythiamine pyrophosphate were similar to cofactor binding site of PfTk. An integrated pharmacophore, co-factor ThDP and substrate fructose-6-pho- sphate, based virtual screening of a small mo- lecule database retrieved eight and thirteen compounds respectively. When screened for their activity against P. falciparum transketolase, one compound in case of ThDP and three compounds in case of fructose-6-phosphate based screening were found active against PfTk. Identification of these novel and chemically diverse inhibitors provides initial leads for optimization of more potent and efficacious drug candidates to treat malarial infection.

Leishmania donovani dipeptidylcarboxypeptidsae (LdDCP), an angiotensin converting enzyme (ACE) related metallopeptidase has been identified and characterized as a putative drug target for antileishmanial chemotherapy. The kinetic parameters for LdDCP with substrate, Hip-His-Leu were determined as, Km, 4 mM and Vmax, 1.173 micromole/ml/min. Inhibition studies revealed that known ACE inhibitors (captopril and bradykinin potentiating peptide; BPP1) were weak inhibitors for LdDCP as compared to human testicular ACE (htACE) with Ki values of 35.8 nM and 3.9 microM, respectively. Three dimensional model of LdDCP was generated based on crystal structure of Escherichia coli DCP (EcDCP) by means of comparative modeling and assessed using PROSAII, PROCHECK and WHATIF. Captopril docking with htACE, LdDCP and EcDCP and analysis of molecular electrostatic potentials (MEP) suggested that the active site domain of three enzymes has several minor but potentially important structural differences. These differences could be exploited for designing selective inhibitor of LdDCP thereby antileishmanial compounds either by denovo drug design or virtual screening of small molecule databases.

The protozoan parasite Leishmania donovani is the causative agent of visceral leishmaniasis. The enzyme pteridine reductase 1 (PTR1) of L. donovani acts as a metabolic bypass for drugs targeting dihydrofolate reductase (DHFR); therefore, for successful antifolate chemotherapy to be developed against Leishmania, it must target both enzyme activities. Leishmania cells overexpressing PTR1 tagged at the N-terminal with green fluorescent protein were established to screen for proprietary dihydropyrimidone (DHPM) derivatives of DHFR specificity synthesised in our laboratory. A cell-permeable molecule with impressive antileishmanial in vitro and in vivo oral activity was identified. Structure activity relationship based on homology model drawn on our recombinant enzyme established the highly selective inhibition of the enzyme by this analogue. It was seen that the leishmanicidal effect of this analogue is triggered by programmed cell death mediated by the loss of plasma membrane integrity as detected by binding of annexin V and propidium iodide (PI), loss of mitochondrial membrane potential culminating in cell cycle arrest at the sub-G0/G1 phase and oligonucleosomal DNA fragmentation. Hence, this DHPM analogue [(4-fluoro-phenyl)-6-methyl-2-thioxo-1, 2, 3, 4-tetrahydropyrimidine-5-carboxylic acid ethyl ester] is a potent antileishmanial agent that merits further pharmacological investigation.

A total of 80 new 2-methyl-6-ureido-4-quinolinamides were synthesized and evaluated for their antimalarial activity. Several analogs elicited the antimalarial effect at MIC of 0.25 mg/mL against the chlooquine-sensitive P. falciparum strain. The IC 50 values of the active compounds were observed to be in ng/mL range and two of the analogs have better IC 50 value than the standard chloroquine. In the in vivo assay against mdr CQ resistant P. yoelii N67/ P. yoelii nigeriensis , however, none of the compound showed complete suppression of parasitemia on day 7. One of the compounds displayed significant antibacterial effect against several strains of bacteria and was many-fold better than the standard drug gentamicin.

InhA, the enoyl acyl carrier protein reductase (EACP reductase) from Mycobacterium tuberculosis, is one of the key enzymes involved in the mycobacterial fatty acid elongation cycle and has been validated as an effective target for the development of anti-microbial agents. We report here, comparative molecular field analysis (CoMFA) studies and subsequent de novo ligand design using the LeapFrog program on pyrrolidine carboxamides, which have been reported as selective inhibitors of EACP reductase from Mycobacterium tuberculosis. The CoMFA model, constructed from the inhibitors used in this study has been successfully used to rationalize the structure-activity relationship of pyrrolidine carboxamides. The CoMFA model produced statistically significant results with cross-validated and conventional correlation coefficients of 0.626 and 0.953 respectively. Further, the predictive ability of CoMFA model was determined using a test set which gave predictive correlation coefficient r 2 pred of 0.880, indicating good predictive power. Finally, Leapfrog was used to propose 13 new pyrrolidine carboxamide analogues, based on the information derived from the CoMFA contour maps. The designed molecules showed better predicted activity using the CoMFA model with respect to the already reported systems; hence suggesting that newly proposed molecules in this series of compounds may be more potent and selective toward EACP reductase inhibition.

We have identified new lead candidates that possess inhibitory activity against Mycobacterium tuberculosis H37Rv chorismate mutase by a ligand-based virtual screening optimized for lead evaluation in combination with in vitro enzymatic assay. The initial virtual screening using a ligand-based pharmacophore model identified 95 compounds from an in-house small molecule database of 15,452 compounds. The obtained hits were further evaluated by molecular docking and 15 compounds were short listed based on docking scores and the other scoring functions and subjected to biological assay. Chorismate mutase activity assays identified four compounds as inhibitors of M. tuberculosis chorismate mutase (MtCM) with low K(i) values. The structural models for these ligands in the chorismate mutase binding site will facilitate medicinal chemistry efforts for lead optimization against this protein.

A series of 3-substituted 1, 4-benzodiazepin-2-ones derived from S and R amino acids were evaluated for their anti-ischemic activity in vitro. Treatment with compounds 7h, 16, 9d, and 17 decreased the apoptotic neuronal number, however increased the neuronal viability. The compounds decreasing apoptosis could protect neurons from the ischemic injury. The difference in the activities of 1,4-benzodiazepin-2-ones derived from S- and R-amino acids is discussed and explained on the basis of molecular modeling studies.