Structural Modelling of Platelet Activating Factor Acetyl Hydrolase in Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei: Implications on Therapeutics for Leishmaniasis, Chagas Disease, and Sleeping Sickness.

Purpose: Leishmaniasis, Chagas disease, and sleeping sickness are caused by protozoa Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei, respectively. Platelet activating factor acetyl hydrolase (PAF-AH) is an inflammatory protein implicated in pathogenesis of these three infections, thereby making them attractive drug targets. Methods: PAF-AH sequences were retrieved from UniProt and aligned using Clustal Omega. Homologous models of parasitic proteins were built based on crystal structure of human PAF-AH and validated using PROCHECK server. Volumes of substrate-binding channel were calculated using the ProteinsPlus program. High throughput virtual screening using Glide program in Schrodinger was done with ZINC drug library against parasitic PAF-AH enzymes. Complexes with best hits were energy-minimized and subjected to 100 ns molecular dynamic simulation and analyzed. Results: PAF-AH enzyme sequences from protozoa Leishmania donovani, Trypanosoma cruzi, Trypanosoma brucei, and human have a minimum of 34% sequence similarity with each other. Corresponding structures show a globular conformation consisting of twisted ß-pleated sheets, flanked by α-helices on either side. Catalytic triad of serine-histidine-aspartate is conserved. Substrate-binding channel residues are conserved to an extent, with a lower channel volume in human as compared to target enzymes. Drug screening resulted in identification of three molecules that had better affinities than the substrate to the target enzymes. These molecules fulfill Lipinski's rules for drug likeness and also bind with less affinity to the human counterpart, thereby establishing a high selective index. Conclusion: Structures of PAF-AH from protozoan parasites and humans belong to the same family of enzymes and have a similar three-dimensional fold. However, they show subtle variations in residue composition, secondary structure composition, substrate-binding channel volume, and conformational stability. These differences result in certain specific molecules being potent inhibitors of the target enzymes while simultaneously having weaker binding to human homologue.

Evaluating the potency of Sulawesi propolis compounds as ACE-2 inhibitors through molecular docking for COVID-19 drug discovery preliminary study.

Coronavirus disease (COVID-19) is a global pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Up to date, there has been no specific cure to treat the disease. Indonesia is one of the countries that is still fighting to control virus transmission. Yet, at the same time, Indonesia has a rich biodiversity of natural medicinal products that potentially become an alternative cure. Thus, this study examined the potency of a natural medicinal product, Sulawesi propolis compounds produced by Tetragonula sapiens, inhibiting angiotensin-converting activity enzyme-2 (ACE-2), a receptor of SARS-CoV-2 in the human body. In this study, molecular docking was done to analyze the docking scores as the representation of binding affinity and the interaction profiles of propolis compounds toward ACE-2. The results illustrated that by considering the docking score and the presence of interaction with targeted sites, five compounds, namely glyasperin A, broussoflavonol F, sulabiroins A, (2S)-5,7-dihydroxy-4'-methoxy-8-prenylflavanone and isorhamnetin are potential to inhibit the binding of ACE-2 and SARS-CoV-2, with the docking score of -10.8, -9.9, -9.5, -9.3 and -9.2 kcal/mol respectively. The docking scores are considered to be more favorable compared to MLN-4760 as a potent inhibitor.

Molecular interaction analysis of Sulawesi propolis compounds with SARS-CoV-2 main protease as preliminary study for COVID-19 drug discovery.

Coronavirus disease 2019 (COVID-19), a respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a global health concern, as the World Health Organization declared this outbreak to be a global pandemic in March 2020. The need for an effective treatment is urgent because the development of an effective vaccine may take years given the complexity of the virus and its rapid mutation. One promising treatment target for COVID-19 is SARS-CoV-2 main protease. Thus, this study was aimed to examine whether Sulawesi propolis compounds produced by Tetragonula sapiens inhibit the enzymatic activity of SARS-CoV-2 main protease. In this study, molecular docking was performed to analyze the interaction profiles of propolis compounds with SARS-CoV-2 main protease. The results illustrated that two compounds, namely glyasperin A and broussoflavonol F, are potential drug candidates for COVID-19 based on their binding affinity of -7.8 kcal/mol and their ability to interact with His41 and Cys145 as catalytic sites. Both compounds also displayed favorable interaction profiles with SARS-CoV-2 main protease with binding similarities compared to inhibitor 13b as positive control 63% and 75% respectively.

Clustering and Sampling of the c-Met Conformational Space: A Computational Drug Discovery Study.

BACKGROUND: c-Met kinase plays a critical role in a myriad of human cancers, and a massive scientific work was devoted to design more potent inhibitors. OBJECTIVE: In this study, 16 molecular dynamics simulations of different complexes of potent c-Met inhibitors with U-shaped binding mode were carried out regarding the dynamic ensembles to design novel potent inhibitors. METHODS: A cluster analysis was performed, and the most representative frame of each complex was subjected to the structure-based pharmacophore screening. The GOLD docking program investigated the interaction energy and pattern of output hits from the virtual screening. The most promising hits with the highest scoring values that showed critical interactions with c-Met were presented for ADME/Tox analysis. RESULTS: The screening yielded 45,324 hits that all of them were subjected to the docking studies and 10 of them with the highest-scoring values having diverse structures were presented for ADME/Tox analyses. CONCLUSION: The results indicated that all the hits shared critical Pi-Pi stacked and hydrogen bond interactions with Tyr1230 and Met1160 respectively.

Expression, Purification and Characterization of a GII.4 Norovirus Protease from Minerva Virus.

BACKGROUND: Noroviruses are the leading cause of acute gastroenteritis worldwide. Norovirus proteases, which are responsible for cleavage of the viral polyprotein, have become an attractive drug target to treat norovirus infections. Genogroup II (GII) noroviruses are responsible for a majority of outbreaks; however, limited data exists regarding GII norovirus proteases. METHODS: We report here successful expression, purification, characterization, and inhibition of the Minerva virus protease (MVpro), a genogroup II genotype 4 (GII.4) norovirus protease. We observed MVpro as both a monomer and dimer in solution through sizeexclusion chromatography. In addition, MVpro cleaves the synthetic substrate mimicking the MVpro NS2/NS3 cleavage site more efficiently than other norovirus proteases such as the Norwalk virus protease (GI.1) and the MD145 protease (GII.4). RESULTS AND CONCLUSION: Compound A, a potent inhibitor of MVpro, is a good starting point for the design of inhibitors to target GII.4 noroviruses. Furthermore, the results presented here will allow for future characterization of MVpro inhibitors as they are synthesized.

Development of potent chemical antituberculosis agents targeting Mycobacterium tuberculosis acetohydroxyacid synthase.

Mycobacterium tuberculosis acetohydroxyacid synthase (MTB-AHAS) has been suggested as a crucial target for antibacterial agents. High-throughput screening of a chemical library was performed to identify potent new inhibitors of MTB-AHAS. Among the 6800 tested compounds, 15 were identified as potent inhibitors, exhibiting >80-90% inhibition of in vitro MTB-AHAS activity at a fixed concentration of 20 µM. Five compounds belonging to the triazolopyrimidine structural class showed greater inhibition potency, with a half-maximum inhibition concentration (IC50 value) in the low micromolar range (0.4-1.24 µM). Furthermore, potent inhibitors demonstrated non-competitive, uncompetitive or mixed-competitive inhibition. Molecular docking experiments with these potent chemicals using a homology model of MTB-AHAS indicated hydrophobic and hydrogen bond interactions with some key herbicide binding site residues with binding energies (ΔG) of -8.04 to -10.68 Kcal/mol, respectively. The binding modes were consistent with inhibition mechanisms, as the chemicals were oriented outside the active site. Importantly, these potent inhibitors demonstrated significant growth inhibition of various clinically isolated multidrug-resistant and extensively drug-resistant M. tuberculosis strains, with 50% minimum inhibitory concentrations (MIC50 values) ranging from 0.2 µg/mL to 0.8 µg/mL, which resemble the MICs of conventional drugs for tuberculosis (isoniazid, 0.1 µg/mL; rifampicin, 0.4 µg/mL). Thus, the identified potent inhibitors show potential as scaffolds for further in vivo studies and might provide an impetus for the development of strong antituberculosis agents targeting MTB-AHAS.

Potent inhibition of cytochrome P450 2B6 by sibutramine in human liver microsomes.

The present study was performed to evaluate the potency and specificity of sibutramine as an inhibitor of the activities of nine human CYP isoforms in liver microsomes. Using a cocktail assay, the effects of sibutramine on specific marker reactions of the nine CYP isoforms were measured in human liver microsomes. Sibutramine showed potent inhibition of CYP2B6-mediated bupropion 6-hydroxylation with an IC50 value of 1.61µM and Ki value of 0.466µM in a competitive manner at microsomal protein concentrations of 0.25mg/ml; this was 3.49-fold more potent than the typical CYP2B6 inhibitor thio-TEPA (Ki=1.59µM). In addition, sibutramine slightly inhibited CYP2C19 activity (Ki=16.6µM, noncompetitive inhibition) and CYP2D6 activity (Ki=15.7µM, noncompetitive inhibition). These observations indicated 35.6- and 33.7-fold decreases in inhibition potency, respectively, compared with that of CYP2B6 by sibutramine. However, no inhibition of CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2D6, or CYP2E1 activities was observed. In addition, the CYP2B6 inhibitory potential of sibutramine was enhanced at a lower microsomal protein concentration of 0.05mg/ml. After 30min preincubation of human liver microsomes with sibutramine in the presence of NADPH, no shift in IC50 was observed in terms of inhibition of the activities of the nine CYPs, suggesting that sibutramine is not a time-dependent inactivator. These observations suggest that sibutramine is a selective and potent inhibitor of CYP2B6 in vitro, whereas inhibition of other CYPs is substantially lower. These in vitro data support the use of sibutramine as a well-known inhibitor of CYP2B6 for routine screening of P450 reversible inhibition when human liver microsomes are used as the enzyme source.