Identification of amentoflavone as a potent SARS-CoV-2 Mpro inhibitor: a combination of computational studies and in vitro biological evaluation.

Small-molecule inhibitors of SARS-CoV-2 Mpro that block the active site pocket of the viral main protease have been considered potential therapeutics for the development of drugs against SARS-CoV-2. Here, we report the identification of amentoflavone (a biflavonoid) through docking-based virtual screening of a library comprised of 231 compounds consisting of flavonoids and isoflavonoids. The docking results were further substantiated through extensive analysis of the data obtained from all-atom 150 ns MD simulation. End-state effective free energy calculations using MM-PBSA calculations further suggested that (Ra)-amentoflavone (C3'-C8''-atropisomer) may show a greater binding affinity towards the Mpro than (Sa)-amentoflavone. In vitro cytotoxicity assay established that amentoflavone showed a high CC50 value indicating much lower toxicity. Further, potent inhibition of the Mpro by amentoflavone was established by studying the effect on HEK293T cells treated with SARS-CoV-2 Mpro expressing plasmid.Communicated by Ramaswamy H. Sarma.

Exploring quantum computational, molecular docking, and molecular dynamics simulation with MMGBSA studies of ethyl-2-amino-4-methyl thiophene-3-carboxylate.

2-aminothiophenes derivative, Ethyl-2-amino-4-methyl thiophene-3-carboxylate (EAMC) has been synthesized, characterized, and investigated quantum chemically. It was experimentally investigated by different spectroscopic methods like- NMR (1H-NMR and 13C-NMR), FT-IR, and UV-Visible. B3LYP method and 6-311++G(d,p) basis set were employed for optimization of molecular structure and calculation of wave numbers of normal modes of vibrations and various other important parameters. Calculated bond lengths and angles were compared with the experimental bond lengths and Bond Angle Parameters. Optimized bond parameters and experimental bond parameters were found in good agreement. Complete potential energy distribution assignments were done successfully by VEDA. The HOMO/LUMO energy gap emphasizes adequate charge transfer happening within the molecule. A study of donor-acceptor interconnections was done via NBO analysis. MEP surface analysis was done to demonstrate charge distribution and reactive areas qualitatively in the molecule. The degree of relative localization of electrons was analyzed via ELF Diagram. The Fukui function analysis showed possible sites for attacks by different substituents. By using the TD-DFT method and PCM solvent model, the UV-Vis spectrum (gas, methanol, DMSO) and the maximum absorption wavelength was computed and compared with experimental data. 3D and 2D intermolecular interactions in the crystal were analyzed via Hirshfeld surface analysis and fingerprint plots reveal that the EAMC crystal was stabilized by H--H/H--H/C--H bond formation. The molecular docking was done with 7 different protein receptors on the molecule to find the best ligand-protein interactions. Molecular dynamic simulations and MMGBSA calculations were also carried out to find out the best binding of the ligand with the protein.Communicated by Ramaswamy H. Sarma.

Evaluation of experimental, computational, molecular docking and dynamic simulation of flucytosine.

Flucytosine (5-fluorocytosine), a fluorine derivative of pyrimidine, has been studied both experimentally and quantum chemically. To obtain the optimized structure, vibrational frequencies and other various parameters, the B3LYP method with a 6-311++G(d,p) basis set was used. Atom-in-molecule theory was used to calculate the binding energies, ellipticity and isosurface projection by electron localization of the molecule (AIM). In addition, the computational results from IR and Raman were compared with the experimental spectra. NBO analysis was used to analyze the donor and acceptor interactions. To know the reactive region of the molecule, the molecular electrostatic potential (MEP) and Fukui functions were determined. The UV-Vis spectrum calculated by the TD-DFT/PCM method was also compared with the experimentally determined spectrum. The HOMO-LUMO energy outcomes proved that there was a good charge exchange occurring within the molecule. With DMSO and MeOH as the solvents, maps of the hole and electron density distribution (EDD and HDD) were produced in an excited state. An electrophilicity index parameter was looked at to theoretically test the bioactivity of the compound. To find the best ligand-protein interactions, molecular docking was also carried out with various receptor proteins. In order to verify the inhibitory potency for the receptor protein complex predicted by docking and molecular dynamic simulation studies, the binding free energy of the receptor protein complex was calculated. Using the MM/GBSA technique, we determined the docked complex's binding free energy. To confirm the molecule's drug similarity, a biological drug similarity investigation was also executed.Communicated by Ramaswamy H. Sarma.

Angle distortion model for predicting enediyne activation towards Bergman cyclization: an alternate to the distance theory.

The kinetics of Bergman cyclization (BC) of enediynes into 1,4-benzene diradicals (also known as p-benzynes) have attracted interest ever since the discovery of natural enediynes which pointed out a surprising reactivity profile difference across enediynes with varying structural architectures. From the analysis of experimental kinetic data, several models were proposed to have a structure-kinetics correlation, out of which, the cd-distance model and the transition state model are the most accepted ones. Recently, Houk et al. introduced a distortion model to explain the regioselectivity of nucleophilic addition to unsymmetrical o-benzynes based on the geometry of the transition state. In the case of BC, since the reaction is endothermic, the transition state geometrically resembles the product structure which implies that in the reaction pathway, the sp-carbons of enediynes are transformed into the trigonal sp2 carbons of the benzenoid product. Thus, greater bending of the interior angles at the proximal alkyne carbons in the enediynes will lead to a lower activation barrier for the BC and hence faster cyclization. This hypothesis has been tested on a series of enediynes including natural product surrogates and the extent of deviation correlates well with the kinetic results. A cut-off value for the average internal proximal angles has been proposed to categorize enediynes as per their reactivity under ambient conditions. We believe that this distortion theory offers an alternative model in designing new unnatural enediynes with desired kinetic stabilities.

Targeting an evolutionarily conserved "E-L-L" motif in the spike protein to develop a small molecule fusion inhibitor against SARS-CoV-2.

As newer variants of SARS-CoV-2 continue to pose major threats to global human health and economy, identifying novel druggable antiviral targets is the key towards sustenance. Here, we identify an evolutionary conserved E-L-L motif present within the HR2 domain of all human and non-human coronavirus spike (S) proteins that play a crucial role in stabilizing the post-fusion six-helix bundle (6-HB) structure and thus, fusion-mediated viral entry. Mutations within this motif reduce the fusogenicity of the S protein without affecting its stability or membrane localization. We found that posaconazole, an FDA-approved drug, binds to this E-L-L motif resulting in effective inhibition of SARS-CoV-2 infection in cells. While posaconazole exhibits high efficacy towards blocking S protein-mediated viral entry, mutations within the E-L-L motif rendered the protein completely resistant to the drug, establishing its specificity towards this motif. Our data demonstrate that posaconazole restricts early stages of infection through specific inhibition of membrane fusion and viral genome release into the host cell and is equally effective towards all major variants of concerns of SARS-CoV-2 including beta, kappa, delta, and omicron. Together, we show that this conserved essential E-L-L motif is an ideal target for the development of prophylactic and therapeutic interventions against SARS-CoV-2.

Targeting an evolutionarily conserved "E-L-L" motif in spike protein to identify a small molecule fusion inhibitor against SARS-CoV-2.

As newer variants of SARS-CoV-2 continue to pose major threats to global human health and economy, identifying novel druggable antiviral targets is the key toward sustenance. Here, we identify an evolutionarily conserved "Ex3Lx6L" ("E-L-L") motif present within the HR2 domain of all human and nonhuman coronavirus spike (S) proteins that play a crucial role in stabilizing its postfusion six-helix bundle (6-HB) structure and thus, fusion-mediated viral entry. Mutations within this motif reduce the fusogenicity of the S protein without affecting its stability or membrane localization. We found that posaconazole, an FDA-approved drug, binds to this "E-L-L" motif and impedes the formation of 6-HB, thus effectively inhibiting SARS-CoV-2 infection in cells. While posaconazole exhibits high efficacy in blocking S protein-mediated viral entry, mutations within the "E-L-L" motif rendered the protein completely resistant to the drug, establishing its specificity toward this motif. Our data demonstrate that posaconazole restricts early stages of infection through specific inhibition of membrane fusion and viral genome release into the host cell and is equally effective toward all major variants of concerns of SARS-CoV-2, including Beta, Kappa, Delta, and Omicron. Together, we show that this conserved essential "E-L-L" motif is an ideal target for the development of prophylactic and therapeutic interventions against SARS-CoV-2.

Sterically hindering the trajectory of nucleophilic attack towards p-benzynes by a properly oriented hydrogen atom: an approach to achieve regioselectivity.

Nucleophilic addition to p-benzynes derived via Bergman cyclization has become a topic of keen interest. Studying the regioselectivity in such addition can reveal important information regarding the parameters controlling such addition. Recently, high regioselectivity has been achieved in nucleophilic addition to a p-benzyne derived from an ortho substituted benzo fused cyclic azaenediyne. Rather than having a freely rotating substitution, a rigid hydrogen atom coming from a suitable naptho fused enediyne and residing in the plane of the p-benzyne ring can offer hindrance to the trajectory of the nucleophile. This can lead to regioselectivity provided the other side remains relatively free of such hindrance. Based on that approach, halide addition to p-benzynes derived from naphtho fused cyclic azaenediynes was studied and a high level of regioselectivity was observed. Steric hindrance to the trajectory of nucleophile by the bay hydrogen was found to be the main cause of such regioselectivity; however, differential electrostatic potential as well as distortions at reactive centres have a minor role in controlling the regioselectivity. The products of such high yielding addition are the halo naphtho tetrahydroisoquinolines.

Fluoroquinolone antibiotics show genotoxic effect through DNA-binding and oxidative damage.

The fluoroquinolones (FQs) are one the most successful class of synthetic antibiotics that primarily target the type II topoisomerases. With a pursuit to evaluate their genotoxicity, the present work established moderate to good DNA-damaging properties of some of the well-known and clinically prescribed fluoroquinolone antibiotics (2nd and 3rd generation). Hypochromic shift in UV-Vis absorption titration, fluorescence quenching in competitive ethidium bromide displacement assay (with calf-thymus DNA) and in-silico studies established DNA-intercalation with binding constants of the order 104. A basic Structure Activity Relationship (SAR) has been derived from the docking results. MTT assay has been also done to evaluate the effect of these antibiotics on cell viability. The expression level of specific DNA-glycosylase enzymes responsible for repairing the oxidized DNA bases are quantified through western blot analysis. The studies revealed that fluoroquinolone antibiotics initiate the genotoxic effect at a concentration of above 50 µg/mL. Recruitment of APE1 and NEIL1 was found to be significantly increased to remove the oxidized nucleobases.

Photochemical Activation of Enediyne Warheads: A Potential Tool for Targeted Antitumor Therapy.

Spatial and temporal control over DNA cleavage by photoactivated enediynes can be complemented by additional factors such as the release of internal strain, chelation, pH changes, intramolecular H-bonds, and substituent effects. This review presents design and reactivity of photoactivated enediynes/enynes and analyses the chemical, biological, and photophysical challenges in their applications.

Chloramphenicol-borate/boronate complex for controlling infections by chloramphenicol-resistant bacteria.

Increasing bacterial resistance to antibiotics is a pressing problem worldwide, with many health organisations prioritizing this issue. Whilst there is a desperate need for new effective antimicrobials, it is also important to understand the mechanisms and epidemiology of the resistant pathogens currently present in the community. Chloramphenicol is one such well known antibiotic which had lost its efficacy due to bacterial resistance. In this paper, we report the design, synthesis, and bio-studies of novel chloramphenicol-borate/boronate derivatives which showed the ability to control the infections caused by chloramphenicol-resistant bacteria. Activity profiling against P. aeruginosa strain EXR1 with catB gene indicated the inability of acetyl transferase to acetylate the chloramphenicol-borate/boronate complex, unlike chloramphenicol. Results obtained from the antimicrobial assays were further rationalized by molecular docking studies. The latter revealed that the probable reason for the enhanced antibacterial activity may be attributed to the change in the binding site of chloramphenicol-borate/boronate with chloramphenicol acetyl transferase (CAT) with respect to chloramphenicol itself. Hemolytic and genotoxic studies established the reduced toxicity of these synthetic derivatives with respect to chloramphenicol.

Use of azidonaphthalimide carboxylic acids as fluorescent templates with a built-in photoreactive group and a flexible linker simplifies protein labeling studies: applications in selective tagging of HCAII and penicillin binding proteins.

This work describes the synthesis of azidonaphthalimide carboxylic acids which act as fluorescent templates with a built-in photoreactive group and a linker thus simplifying the design of protein labeling agents. These were separately connected to selectivity hands like a sulfonamide and ampicillin for successful labeling/detection of HCAII and PBPs, respectively.

Asymmetric Garratt-Braverman cyclization: a route to axially chiral aryl naphthalene-amino acid hybrids.

We report the first example of a highly diastereoselective Garratt-Braverman cyclization leading to the synthesis of chiral aryl naphthalene-amino acid hybrids in excellent yields. The stereogenecity in the amino acid has induced high diastereoselectivity for the reaction. Computations based on density functional theory indicated a lower activation free energy barrier for the M isomer as compared to that for the P diastereomer (ΔΔG = 3.48 kcal/mol). Comparison of the recorded CD spectrum of the product with the calculated one also supported the preferential formation of the M diastereomer.