Evaluating the use of absolute binding free energy in the fragment optimization process.

Key to the fragment optimization process is the need to accurately capture the changes in affinity that are associated with a given set of chemical modifications. Due to the weakly binding nature of fragments, this has proven to be a challenging task, despite recent advancements in leveraging experimental and computational methods. In this work, we evaluate the use of Absolute Binding Free Energy (ABFE) calculations in guiding fragment optimization decisions, retrospectively calculating binding free energies for 59 ligands across 4 fragment elaboration campaigns. We first demonstrate that ABFEs can be used to accurately rank fragment-sized binders with an overall Spearman’s r of 0.89 and a Kendall τ of 0.67, although often deviating from experiment in absolute free energy values with an RMSE of 2.75 kcal/mol. We then also show that in several cases, retrospective fragment optimization decisions can be supported by the ABFE calculations. Cases that were not supported were often limited by large uncertainties in the free energy estimates, however generally the right direction in ΔΔG is still observed. Comparing against cheaper endpoint methods, namely Nwat-MM/GBSA, we find that ABFEs offer better outcomes in ranking binders, improving correlation metrics, although a similar confidence in retrospective synthetic decisions is achieved. Our results indicate that ABFE calculations are currently at the level of accuracy that can be usefully employed to gauge which fragment elaborations are likely to offer the best gains in affinity.

Identification of Some Dietary Flavonoids as Potential Inhibitors of TMPRSS2 Through Protein-ligand Interaction Studies and Binding Free Energy Calculations

The alarming increase in COVID-19 cases and deaths calls for an urgent cost-effective pharmacological approach. Here, we examine the inhibitory activity of a group of dietary bioactive flavonoids against the human protease TMPRSS2, which plays a major role in SARS CoV-2 viral entry. After the molecular docking studies of a large number of flavonoids, four compounds with high binding scores were selected and studied in detail. The binding affinities of these four ligands, Amentoflavone, Narirutin, Eriocitrin, and Naringin, at the active site of TMPRSS2 target were investigated using MD simulations followed by MM-PBSA binding energy calculations. From the studies, a number of significant hydrophobic and hydrogen bonding interactions between the ligands and binding site amino residues of TMPRSS2 are identified which showcase their excellent inhibitory activity against TMPRSS2. Among these ligands, Amentoflavone and Narirutin showed MM-PBSA binding energy values of -155.48 and -138.13 kJ/mol respectively. Our previous studies of the inhibitory activity of these compounds against main protease of SARS-COV2 and the present study on TMPRSS2 strongly highlighted that Amentoflavone and Naringin can exhibit promising multi-target activity against SARS-CoV-2. Moreover, due to their wide availability, no side effects and low cost, these compounds could be recommended as dietary supplements for COVID patients or for the development of SARS-CoV-2 treatments.

In Silico-Based Discovery of Natural Anthraquinones with Potential against Multidrug-Resistant E. coli

E. coli is a Gram-negative bacterium that causes different human infections. Additionally, it resists common antibiotics due to its outer protective membrane. Natural products have been proven to be efficient antibiotics. However, plant natural products are far less explored in this regard. Accordingly, over 16,000 structures covering almost all African medicinal plants in AfroDb in a structural-based virtual screening were used to find efficient anti-E. coli candidates. These drug-like structures were docked into the active sites of two important molecular targets (i.e., E. coli’s Ddl-B and Gyr-B). The top-scoring hits (i.e., got docking scores < −10 kcal/mol) produced in the initial virtual screening (0.15% of the database structures for Ddl-B and 0.17% of the database structures for Gyr-B in the database) were further refined using molecular dynamic simulation-based binding free energy (ΔG) calculation. Anthraquinones were found to prevail among the retrieved hits. Accordingly, readily available anthraquinone derivatives (10 hits) were selected, prepared, and tested in vitro against Ddl-B, Gyr-B, multidrug-resistant (MDR) E. coli, MRSA, and VRSA. A number of the tested derivatives demonstrated strong micromolar enzyme inhibition and antibacterial activity against E. coli, MRSA, and VRSA, with MIC values ranging from 2 to 64 µg/mL. Moreover, both E. coli’s Ddl-B and Gyr-B were inhibited by emodin and chrysophanol with IC50 values comparable to the reference inhibitors (IC50 = 216 ± 5.6, 236 ± 8.9 and 0.81 ± 0.3, 1.5 ± 0.5 µM for Ddl-B and Gyr-B, respectively). All of the active antibacterial anthraquinone hits showed low to moderate cellular cytotoxicity (CC50 > 50 µM) against human normal fibroblasts (WI-38). Furthermore, molecular dynamic simulation (MDS) experiments were carried out to reveal the binding modes of these inhibitors inside the active site of each enzyme. The findings presented in this study are regarded as a significant step toward developing novel antibacterial agents against MDR strains.

Digalloyl Glycoside: A Potential Inhibitor of Trypanosomal PFK from Euphorbia abyssinica J.F. Gmel

Human African trypanosomiasis is an endemic infectious disease caused by Trypanosoma brucei via the bite of tsetse-fly. Most of the drugs used for the treatment, e.g., Suramin, have shown several problems, including the high level of toxicity. Accordingly, the discovery of anti-trypanosomal drugs from natural sources has become an urgent requirement. In our previous study on the anti-trypanosomal potential of Euphorbia species, Euphorbia abyssinica displayed significant anti-trypanosomal activity. Therefore, a phytochemical investigation of the methanolic extract of E. abyssinica was carried out. Twelve compounds, including two triterpenes (1, 2); one sterol-glucoside (4); three ellagic acid derivatives (3, 9, 11); three gallic acid derivatives (5, 6, 10); and three flavonoids (7, 8, 12), were isolated. The structures of isolated compounds were determined through different spectroscopic techniques. Compound (10) was obtained for the first time from genus Euphorbia while all other compounds except compound (4), were firstly reported in E. abyssinica. Consequently, an in silico study was used to estimate the anti-trypanosomal activity of the isolated compounds. Several compounds displayed interesting activity where 1,6-di-O-galloyl-d-glucose (10) appeared as the most potent inhibitor of trypanosomal phosphofructokinase (PFK). Moreover, molecular dynamics (MD) simulations and ADMET calculations were performed for 1,6-di-O-galloyl-d-glucose. In conclusion, 1,6-di-O-galloyl-d-glucose revealed high binding free energy as well as desirable molecular dynamics and pharmacokinetic properties; therefore, it could be suggested for further in vitro and in vivo studies for trypanosomiasis.

Combinatorial Virtual Screening Revealed a Novel Scaffold for TNKS Inhibition to Combat Colorectal Cancer

The abnormal Wnt signaling pathway leads to a high expression of β-catenin, which causes several types of cancer, particularly colorectal cancer (CRC). The inhibition of tankyrase (TNKS) activity can reduce cancer cell growth, invasion, and resistance to treatment by blocking the Wnt signaling pathway. A pharmacophore search and pharmacophore docking were performed to identify potential TNKS inhibitors in the training databases. The weighted MM/PBSA binding free energy of the docking model was calculated to rank the databases. The reranked results indicated that 26.98% of TNKS inhibitors that were present in the top 5% of compounds in the database and near an ideal value ranked 28.57%. The National Cancer Institute database was selected for formal virtual screening, and 11 potential TNKS inhibitors were identified. An enzyme-based experiment was performed to demonstrate that of the 11 potential TNKS inhibitors, NSC295092 and NSC319963 had the most potential. Finally, Wnt pathway analysis was performed through a cell-based assay, which indicated that NSC319963 is the most likely TNKS inhibitor (pIC50 = 5.59). The antiproliferation assay demonstrated that NSC319963 can decrease colorectal cancer cell growth; therefore, the proposed method successfully identified a novel TNKS inhibitor that can alleviate CRC.

Structural and computational insights into the SARS-CoV-2 Omicron RBD-ACE2 interaction

Since SARS-CoV-2 Omicron variant (B.1.1.529) was reported in November 2021, it has quickly spread to many countries and outcompeted the globally dominant Delta variant in several countries. The Omicron variant contains the largest number of mutations to date, with 32 mutations located at spike (S) glycoprotein, which raised great concern for its enhanced viral fitness and immune escape[1-4]. In this study, we reported the crystal structure of the receptor binding domain (RBD) of Omicron variant S glycoprotein bound to human ACE2 at a resolution of 2.6 Å. Structural comparison, molecular dynamics simulation and binding free energy calculation collectively identified four key mutations (S477N, G496S, Q498R and N501Y) for the enhanced binding of ACE2 by the Omicron RBD compared to the WT RBD. Representative states of the WT and Omicron RBD-ACE2 systems were identified by Markov State Model, which provides a dynamic explanation for the enhanced binding of Omicron RBD. The effects of the mutations in the RBD for antibody recognition were analyzed, especially for the S371L/S373P/S375F substitutions significantly changing the local conformation of the residing loop to deactivate several class IV neutralizing antibodies.

Molecular Dynamics Free Energy Simulation study to Investigate Binding Pattern of Isoliquiritigenin as PPAR𝛾 Agonist

Phytochemicals are rich source of bioactive constituents and can be used as another alternative to currently used drugs for diseases like Diabetes mellitus. The potential of Isoliquiritigenin (a constituent of Pterocarpus marsupium) as PPAR𝛾 agonist was evaluated by in silico technique. Autodock results showed that Tyr327, and Tyr473 of the PPARγ forms H-bonds with Isoliquiritigenin (binding energy of -7.46 kcal/mol) and Troglitazone (known drug) showed H bond with Tyr327, Ser289, with binding energy of -11.01 kcal/mol. Isoliquiritigenin, binding energy in Extra precision (XP) was -6.74 kcal/mol while Troglitazone docking, gave binding energy in XP mode as -9.59 kcal/mol. The best Induced fit docking (IFD) score of the optimised PPARγ- Isoliquiritigenin complexes was -9.39 Kcal/mol. The important residues in IFD forming H bond were Cys 285, Arg 288, Tyr 327 and Leu 340. The post docking MM/GBSA free energy for PPARγ with Isoliquiritigenin and Troglitazone was -49.29 and -71.48 Kcal/mol respectively. Binding interaction in MD simulation and Principal Component Analysis studies revealed stable binding throughout 100 ns simulation. Post Simulation MM/PBSA free energy was calculated. The results indicated that compound possessed a negative binding free energy with -114.37KJ/mol. It was observed that van der Waals, electrostatic interactions and non-polar solvation energy negatively contributed to the total interaction energy while only polar solvation energy positively contributed to total free binding energy. The Isoliquiritigenin fulfils the criteria of drug-likeness property. Thus, study presents a systematic analysis on molecular mechanism of action of Isoliquiritigenin as PPARγ agonist in controlling Diabetes mellitus.

Computational study of the furin cleavage domain of SARS-CoV-2: delta binds strongest of extant variants

We demonstrate that AlphaFold and AlphaFold Multimer, implemented within the ColabFold suite, can accurately predict the structures of the furin enzyme with known six residue inhibitory peptides. Noting the similarity of the peptide inhibitors to polybasic furin cleavage domain insertion region of the SARS-CoV-2, which begins at P681, we implement this approach to study the wild type furin cleavage domain for the virus and several mutants. We introduce mutations in silico for alpha, omicron, and delta variants, for several sequences which have been rarely observed, for sequences which have not yet been observed, for other coronaviruses (NL63, OC43, HUK1a, HUK1b, MERS, and 229E), and for the H5N1 flu. We show that interfacial hydrogen bonds between the furin cleavage domain and furin are a good measure of binding strength that correlate well with endpoint binding free energy estimates, and conclude that among all candidate viral sequences studied, delta is near the very top binding strength within statistical accuracy. However, the binding strength of several rare sequences match delta within statistical accuracy. We find that the furin S1 pocket is optimized for binding arginine as opposed to lysine. This residue, typically at sequence position five, contains the most hydrogen bonds to the furin, and hydrogen bond count for just this residue shows a strong positive correlation with the overall hydrogen bond count . We demonstrate that the root mean square backbone C-alpha fluctuation of the first residue in the furin cleavage domain has a strong negative correlation with the interfacial hydrogen bond count. We show by considering the variation with the number of basic residues that the maximum mean number of interfacial hydrogen bonds expected is 15.7 at 4 basic residues.