Venkatesan Suryanarayanan

Assessment of dual inhibition property of newly discovered inhibitors against PCAF and GCN5 through in silico screening, molecular dynamics simulation and DFT approach.

Abstract p300/CBP-associated factor (PCAF) is one among the histone acetyltransferase (HAT) family enzymes. It is involved in the regulation of transcription by modifying the chromatin structure indirectly through the acetylation of histones. It has been emerged as a promising drug target for various types of cancer. A four-point pharmacophore with two hydrogen bond acceptor, one aromatic ring and one hydrophobic feature, was generated for six highly active isothiazolone derivatives as PCAF inhibitors in order to elucidate their anticancer activity. The generated pharmacophore was used for screening three different databases such as Maybridge, Life Chemicals and Chembridge databases. The screened compounds were further filtered through docking studies. Then the compounds were further carried for ADME prediction. The best three compounds BTB09406, F1418-0051 and F1880-1727 were docked to GCN5 to explore the dual inhibitory properties. The conformational stability of the protein–ligand complexes were analyzed through molecular dynamics simulation. Three best compounds were finally went through electronic structure analysis using density functional theory (DFT) at B3LYP/6-31**G level to understand their molecular reactivity. The results obtained from this study exploit that the three best compounds (BTB09406, F1418-0051 and F1880-1727) were found to have more potent and dual inhibitory properties.

Shape and pharmacophore-based virtual screening to identify potential cytochrome P450 sterol 14α-demethylase inhibitors.

Abstract Sterol 14α-demethylase (CYP51) is a cytochrome P450 heme thiolate containing enzyme involved in biosynthesis of membrane sterols, including sterol in animals, ergosterol in fungi, and a variety of C24-modified sterols in plants and protozoa. Several clinical drugs have been developed to reduce the impact of fungal diseases, but their clinical uses have been limited by the emergence of drug resistance and insufficiencies in their antifungal activity. Therefore, in order to identify potential CYP51 inhibitors, we have implemented a virtual screening (VS) protocol by using both phase shape and pharmacophore model (AHHRR) against Asinex, ChemBridge and Maybridge databases. A filtering protocol, including Lipinski filter, number of rotatable bonds and different precisions of molecular docking was applied in hits selection. The results indicated that both shape-based and pharmacophore-based screening yielded the best result with potential inhibitors. The searched compounds were also evaluated with ADME properties, which show excellent pharmacokinetic properties under the acceptable range. We identified potential CYP51 inhibitors for further investigation, they could also be employed to design ligands with enhanced inhibitory potencies and to predict the potencies of analogs to guide synthesis/or prepare synthetic antifungal analogs against CYP51.

A three-dimensional chemical phase pharmacophore mapping, QSAR modelling and electronic feature analysis of benzofuran salicylic acid derivatives as LYP inhibitors

Lymphoid tyrosine phosphatase (LYP), encoded by the PTPN22 gene, has a critical negative regulatory role in T-cell antigen receptor (TCR) and emerged as a promising drug target for human autoimmune diseases. A five-point pharmacophore with two hydrogen bond acceptors, one hydrogen bond donor and two aromatic ring features was generated for a series of benzofuran salicylic acid derivatives as LYP inhibitors in order to elucidate their anti-autoimmune activity. The generated pharmacophore yielded a significant 3D-QSAR model with r2 of 0.9146 for a training set of 27 compounds. The model also showed excellent predictive power with Q2 of 0.7068 for a test set of eight compounds. The investigation of the 3D-QSAR model has revealed the structural insights which could lead to more potent analogues. The most active and inactive compounds were further subjected to electronic structure analysis using density functional theory (DFT) at B3LYP/3−21∗G level to support the 3D-QSAR predictions. The results obtained from this study are expected to be useful in the proficient design and development of benzofuran salicylic acid derivatives as inhibitors of LYP.

Targeting the cyclin-binding groove site to inhibit the catalytic activity of CDK2/cyclin A complex using p27 KIP1 -derived peptidomimetic inhibitors

Functionally activated cyclin-dependent kinase 2 (CDK2)/cyclin A complex has been validated as an interesting therapeutic target to develop the efficient antineoplastic drug based on the cell cycle arrest. Cyclin A binds to CDK2 and activates the kinases as well as recruits the substrate and inhibitors using a hydrophobic cyclin-binding groove (CBG). Blocking the cyclin substrate recruitment on CBG is an alternative approach to override the specificity hurdle of the currently available ATP site targeting CDK2 inhibitors. Greater understanding of the interaction of CDK2/cyclin A complex with p27 (negative regulator) reveals that the Leu-Phe-Gly (LFG) motif region of p27 binds with the CBG site of cyclin A to arrest the malignant cell proliferation that induces apoptosis. In the present study, Replacement with Partial Ligand Alternatives through Computational Enrichment (REPLACE) drug design strategies have been applied to acquire LFG peptide-derived peptidomimetics library. The peptidomimetics function is equivalent with respect to substrate p27 protein fashion but does not act as an ATP antagonist. The combined approach of molecular docking, molecular dynamics (MD), and molecular electrostatic potential and ADME/T prediction were carried out to evaluate the peptidomimetics. Resultant interaction and electrostatic potential maps suggested that smaller substituent is desirable at the position of phenyl ring to interact with Trp217, Arg250, and Gln254 residues in the active site. The best docked poses were refined by the MD simulations which resulted in conformational changes. After equilibration, the structure of the peptidomimetic and receptor complex was stable. The results revealed that the various substrate protein-derived peptidomimetics could serve as perfect leads against CDK2 protein.

Investigating the folding pathway and substrate induced conformational changes in B. malayi Guanylate kinase.

Guanylate kinase is one of the key enzymes in nucleotide biosynthesis. The study highlights the structural and functional properties of Brugia malayi Guanylate kinase (BmGK) in the presence of chemical denaturants. An inactive, partially unfolded, dimeric intermediate was observed at 1-2M urea while GdnCl unfolding showed monomer molten globule like intermediate at 0.8-1.0M. The results also illustrate the protective role of substrates in maintaining the integrity of the enzyme. The thermo stability of protein was found to be significantly enhanced in the presence of the substrates. Furthermore, binding of the substrates, GMP and ATP to BmGK changed its GdnCl induced unfolding pattern. Docking and molecular dynamic simulation performed for native BmGK, BmGK bound to GMP and GMP+ATP showed change in the fluctuation in the region between 130 and 150 residues. Arg134 lost its interaction with GMP and Arg145 interaction shifted to ATP after 40ns simulation upon binding of ATP to BmGK-GMP complex. We, thus, propose the importance of specific rearrangements contributed by binding of substrates which participate in the overall stability of the protein. The work here emphasizes on detailed biophysical characterization of BmGK along with the significant role of substrates in modulating the structural and functional properties of BmGK.

Dissecting the structure–function relationship in lysozyme domain of mycobacteriophage D29‐encoded peptidoglycan hydrolase

Most bacteriophages rapidly infect and kill bacteria and, therefore, qualify as the next generation therapeutics for rapidly emerging drug‐resistant bacteria such as Mycobacterium tuberculosis. We have previously characterized the mycobacteriophage D29‐generated endolysin, Lysin A, for its activity against mycobacteria. Here, we present a detailed characterization of the lysozyme domain (LD) of D29 Lysin A that hydrolyzes peptidoglycan of both gram‐positive and gram‐negative bacteria with high potency. By characterizing an exhaustive LD protein variant library, we have identified critical residues important for LD activity and stability. We further complement our in vitro experiments with detailed in silico investigations. We present LD as a potent candidate for developing phage‐based broad‐spectrum therapeutics.

Exploration of New and Potent Lead Molecules Against CAAX Prenyl Protease I of Leishmania donovani Through Pharmacophore Based Virtual Screening Approach

AIM AND OBJECTIVE Visceral leishmaniasis is a deadly disease left untreated in over 95% of cases. It is characterized by irregular bouts of fever, weight loss, enlargement of the spleen and liver, and anemia. It is highly endemic in the Indian subcontinent. CAAX prenyl protease I of Leishmania donovani is one of the important targets regulating the post translational modification process. Hence identifying potent drug candidate against the target is essential. This study mainly focuses on developing new and potent inhibitors against CAXX prenyl protease I of Leishmania donovani. MATERIALS AND METHODS Pharmacophore based virtual screening was carried out using derivatives of bi-substrate analog farnesyl transferase inhibitors reported against CAAX prenyl proteases I. On the basis of ligand based pharmacophore model we have obtained 5 point pharmacophore AAADR with three hydrogen bond acceptors (A), one hydrogen bond donor (D) and one aromatic ring. The newly identified hits through pharmacophore model were further docked into the active site of the modeled protein. To get further insights of protein ligand interaction we have performed induced fit docking followed by molecular dynamics simulations. The DFT analysis depicts the electronic structure properties of the compounds. These results can be useful for the development of novel and potent CAAX prenyl protease I inhibitors. RESULTS Initially, we have obtained a large number of newly identified hits by screening four different databases further docked into the active site of the protein and 20 compounds were selected on the basis of docking score. Perhaps Induced fit docking was performed to infer protein ligand interaction in a dynamic state and top 5 compounds 7118044, 7806909, LEG12866807, 9208535, SYN 19867403 were found to have good protein ligand interactions with key amino acid residues such as Glu287, His290 and additional interaction like Ile197, Asn209 Tyr253, Phe254, Gly256, Tyr266 with better binding energy -59.794 Kcal/mol, -66.305 Kcal/mol, -70.467 Kcal/mol, -82.474 Kcal/mol, -64.045. The predicted ADME properties are in desirable range and the HOMO/LUMO gap clearly indicates the electrons behavior of the ligands. Molecular dynamics simulations of the protein ligand complex for 20 ns clearly depicts the compounds are stable throughout the simulation time. CONCLUSION The newly identified hits through pharmacophore based screening could be used as potential CAAX prenyl protease I inhibitors of Leishmania donovani.

Explicit Drug Re-positioning: Predicting Novel Drug–Target Interactions of the Shelved Molecules with QM/MM Based Approaches

With the demand to enhance the speed of the drug discovery process there has been an increased usage of computational approaches in drug discovery studies. However because of their probabilistic outcomes, the challenge is to exactly mimic the natural environment which can provide the exact charge polarization effect while estimating the binding energy between protein and ligand. There has been a large number of scoring functions from simple one to the complex one available for estimating binding energy. The quantum mechanics/molecular mechanics (QM/MM) hybrid approach has been the preferred choice of interest since last decade for modeling reactions in biomolecular systems. The application of QM/MM approach has been expanded right from rescoring the already known complexes and depicting the correct position of some novel molecule to ranking a large number of molecules. It is expected that the application of QM/MM-based scoring will grow in all areas of drug discovery. However, the most promising area will be its application in repositioning, that is, assigning novel functions or targets to the already existing drugs, as this would stop the rising attrition rates as well as reduce the overall time and cost of drug discovery procedure.

Vitexin inhibits Aβ25-35 induced toxicity in Neuro-2a cells by augmenting Nrf-2/HO-1 dependent antioxidant pathway and regulating lipid homeostasis by the activation of LXR-α

Amyloid beta (Aβ) formation is one of the neuropathological hallmarks of Alzheimers disease (AD), which induces the generation of reactive oxygen species (ROS), further leading to the alteration of several signalling pathways. In the present study, vitexin has been evaluated for its neuroprotective activity against Aβ25-35 induced toxicity in Neuro-2a cells. Results of cell free studies indicated that vitexin significantly inhibited the aggregation of Aβ25-35. Studies in Neuro-2a cells revealed that Aβ25-35 significantly affected the cell viability by inducing ROS mediated toxicity and apoptosis. However, pre-treatment of Neuro-2a cells with vitexin (50 μM) significantly restored the cell viability up to 92.86 ± 5.57%. Vitexin has been found to inhibit the production of free radicals and suppress ROS mediated lipid peroxidation, protein oxidation and loss of membrane potential. Also, vitexin modulated the expression of genes involved in antioxidant response mechanisms (Nrf-2, HO-1), cholesterol metabolism (LXR-α, APOE, ABCA-1, Seladin-1), and endoplasmic reticulum stress (Grp78, Gadd153) to offer neuroprotection. Aβ25-35 induced caspase-3 activation, and Bax protein expression was also found to be significantly inhibited by vitexin. Taken together, our results indicate that vitexin offers neuroprotection to cells in part via augmenting the antioxidant mechanisms, maintaining lipid homeostasis and inhibiting apoptosis induced by Aβ.

β-Sitosterol targets Trx/Trx1 reductase to induce apoptosis in A549 cells via ROS mediated mitochondrial dysregulation and p53 activation

Abstractβ-Sitosterol (BS), a major bioactive constituent present in plants and vegetables has shown potent anticancer effect against many human cancer cells, but the underlying mechanism remain elusive on NSCLC cancers. We found that BS significantly inhibited the growth of A549 cells without harming normal human lung and PBMC cells. Further, BS treatment triggered apoptosis via ROS mediated mitochondrial dysregulation as evidenced by caspase-3 & 9 activation, Annexin-V/PI positive cells, PARP inactivation, loss of MMP, Bcl-2-Bax ratio alteration and cytochrome c release. Moreover, generation of ROS species and subsequent DNA stand break were found upon BS treatment which was reversed by addition of ROS scavenger (NAC). Indeed BS treatment increased p53 expression and its phosphorylation at Ser15, while silencing the p53 expression by pifithrin-α, BS induced apoptosis was reduced in A549 cells. Furthermore, BS induced apoptosis was also observed in NCI-H460 cells (p53 wild) but not in the NCI-H23 cells (p53 mutant). Down-regulation of Trx/Trx1 reductase contributed to the BS induced ROS accumulation and mitochondrial mediated apoptotic cell death in A549 and NCI-H460 cells. Taken together, our findings provide evidence for the novel anti-cancer mechanism of BS which could be developed as a promising chemotherapeutic drug against NSCLC cancers.