scholarly journals Enhancing Paraoxon Binding to Organophosphorus Hydrolase Active Site

2021 ◽  
Author(s):  
Léa El Khoury ◽  
David Mobley ◽  
Dongmei Ye ◽  
Susan Rempe
Keyword(s):  
Active Site ◽  
Binding Free Energy ◽  
Hot Spot ◽  
Circulatory System ◽  
Kinetic Studies ◽  
Md Simulations ◽  
Binding Mode ◽  
Hydrolytic Activity ◽  
Free Energy Calculations ◽  
Organophosphorus Hydrolase

<p>Organophosphorus (OP) compounds are among the most toxic of chemical substances and widely used as insecticides, pesticides, and chemical warfare agents. The most important enzyme inhibited by OP compounds is acetylcholinesterase (AChe). Inactivation of AChe function results in the accumulation of neurotransmitter, leading to death due to serious respiratory disorders. Organophosphorus hydrolase (OPH), also called phosphotriesterase, is a homo-dimeric metalloenzyme that can hydrolyze various OP agents in the circulatory system, resulting in products that are generally of reduced toxicity. The best OPH substrate found to date is the insecticide diethyl p-nitrophenyl phosphate (paraoxon). Most structural and kinetic studies assume that the binding orientation of paraoxon is identical to that of diethyl 4-methylbenzylphosphonate, which is the only substrate analog co-crystallized with OPH. In the current work, we used a combined docking and molecular dynamics (MD) approach to predict the likely binding mode of paraoxon in the OPH active site. We identified a potential binding mode of paraoxon that does not match the binding mode of diethyl 4-methylbenzylphosphonate. Then, we used the predicted binding mode to run MD simulations on the wild type (WT) OPH complexed with paraoxon, and OPH mutants complexed with paraoxon. Additionally, we identified 3 hot-spot residues (D253, H254, and I255) involved in the stability of the OPH active site. To further assess these predictions, we then experimentally assayed single and double mutants involving these residues (D253E, H254S, I255S, D253E-H254R and D253E-I255G) for hydrolytic activity against paraoxon. Computational structural analysis of protein-substrate dynamics shows different hydrogen bonding profiles for mutants involving D253 (D253E, D253E-H254R, and D253E-I255G) compared to WT OPH. Additionally, the binding free energy calculations and the experimental kinetics (particularly, <i>k</i><sub>cat</sub> and <i>K<sub>M</sub></i>) of the reactions between each OPH mutant and paraoxon show that mutated forms D253E, D253E-H254R, and D253E-I255G exhibit enhanced activity over WT OPH. Interestingly, our experimental results show that the activity of the double mutant D253E-H254R increased by 19-fold compared to WT OPH.</p>

scholarly journals Enhancing Paraoxon Binding to Organophosphorus Hydrolase Active Site

2021 ◽  
Author(s):  
Léa El Khoury ◽  
David Mobley ◽  
Dongmei Ye ◽  
Susan Rempe
Keyword(s):  
Active Site ◽  
Binding Free Energy ◽  
Hot Spot ◽  
Circulatory System ◽  
Kinetic Studies ◽  
Md Simulations ◽  
Binding Mode ◽  
Hydrolytic Activity ◽  
Free Energy Calculations ◽  
Organophosphorus Hydrolase

<p>Organophosphorus (OP) compounds are among the most toxic of chemical substances and widely used as insecticides, pesticides, and chemical warfare agents. The most important enzyme inhibited by OP compounds is acetylcholinesterase (AChe). Inactivation of AChe function results in the accumulation of neurotransmitter, leading to death due to serious respiratory disorders. Organophosphorus hydrolase (OPH), also called phosphotriesterase, is a homo-dimeric metalloenzyme that can hydrolyze various OP agents in the circulatory system, resulting in products that are generally of reduced toxicity. The best OPH substrate found to date is the insecticide diethyl p-nitrophenyl phosphate (paraoxon). Most structural and kinetic studies assume that the binding orientation of paraoxon is identical to that of diethyl 4-methylbenzylphosphonate, which is the only substrate analog co-crystallized with OPH. In the current work, we used a combined docking and molecular dynamics (MD) approach to predict the likely binding mode of paraoxon in the OPH active site. We identified a potential binding mode of paraoxon that does not match the binding mode of diethyl 4-methylbenzylphosphonate. Then, we used the predicted binding mode to run MD simulations on the wild type (WT) OPH complexed with paraoxon, and OPH mutants complexed with paraoxon. Additionally, we identified 3 hot-spot residues (D253, H254, and I255) involved in the stability of the OPH active site. To further assess these predictions, we then experimentally assayed single and double mutants involving these residues (D253E, H254S, I255S, D253E-H254R and D253E-I255G) for hydrolytic activity against paraoxon. Computational structural analysis of protein-substrate dynamics shows different hydrogen bonding profiles for mutants involving D253 (D253E, D253E-H254R, and D253E-I255G) compared to WT OPH. Additionally, the binding free energy calculations and the experimental kinetics (particularly, <i>k</i><sub>cat</sub> and <i>K<sub>M</sub></i>) of the reactions between each OPH mutant and paraoxon show that mutated forms D253E, D253E-H254R, and D253E-I255G exhibit enhanced activity over WT OPH. Interestingly, our experimental results show that the activity of the double mutant D253E-H254R increased by 19-fold compared to WT OPH.</p>

scholarly journals Enhancing Paraoxon Binding to Organophosphorus Hydrolase Active Site

2021 ◽  
Vol 22 (23) ◽  
pp. 12624
Author(s):  
Léa El Khoury ◽  
David L. Mobley ◽  
Dongmei Ye ◽  
Susan B. Rempe
Keyword(s):  
Binding Affinity ◽  
Active Site ◽  
Binding Free Energy ◽  
Hot Spot ◽  
Substrate Binding ◽  
Kinetic Studies ◽  
Md Simulations ◽  
Binding Mode ◽  
Free Energy Calculations ◽  
Organophosphorus Hydrolase

Organophosphorus hydrolase (OPH) is a metalloenzyme that can hydrolyze organophosphorus agents resulting in products that are generally of reduced toxicity. The best OPH substrate found to date is diethyl p-nitrophenyl phosphate (paraoxon). Most structural and kinetic studies assume that the binding orientation of paraoxon is identical to that of diethyl 4-methylbenzylphosphonate, which is the only substrate analog co-crystallized with OPH. In the current work, we used a combined docking and molecular dynamics (MD) approach to predict the likely binding mode of paraoxon. Then, we used the predicted binding mode to run MD simulations on the wild type (WT) OPH complexed with paraoxon, and OPH mutants complexed with paraoxon. Additionally, we identified three hot-spot residues (D253, H254, and I255) involved in the stability of the OPH active site. We then experimentally assayed single and double mutants involving these residues for paraoxon binding affinity. The binding free energy calculations and the experimental kinetics of the reactions between each OPH mutant and paraoxon show that mutated forms D253E, D253E-H254R, and D253E-I255G exhibit enhanced substrate binding affinity over WT OPH. Interestingly, our experimental results show that the substrate binding affinity of the double mutant D253E-H254R increased by 19-fold compared to WT OPH.

Targeting Peptidyl-prolyl Cis-trans Isomerase NIMA-interacting 1: A Structure-based Virtual Screening Approach to Find Novel Inhibitors

2020 ◽  
Vol 16 (5) ◽  
pp. 605-617 ◽  
Author(s):  
Kauê Santana da Costa ◽  
João M. Galúcio ◽  
Deivid Almeida de Jesus ◽  
Guelber Cardoso Gomes ◽  
Anderson Henrique Lima e Lima ◽  
...  
Keyword(s):  
Inhibitory Activity ◽  
Binding Free Energy ◽  
Md Simulations ◽  
Binding Mode ◽  
Free Energy Calculations ◽  
Pentacyclic Triterpenoids ◽  
Substrate Peptide ◽  
Molecular Mechanism Of Action ◽  
Structural Insights ◽  
Virtual Screening Approach

Background : Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1) is an enzyme that isomerizes phosphorylated serine or threonine motifs adjacent to proline residues. Pin1 has important roles in several cellular signaling pathways, consequently impacting the development of multiple types of cancers. Methods: Based on the previously reported inhibitory activity of pentacyclic triterpenoids isolated from the gum resin of Boswellia genus against Pin1, we designed a computational experiment using molecular docking, pharmacophore filtering, and structural clustering allied to molecular dynamics (MD) simulations and binding free energy calculations to explore the inhibitory activity of new triterpenoids against Pin1 structure. Results: Here, we report different computational evidence that triterpenoids from neem (Azadirachta indica A. Juss), such as 6-deacetylnimbinene, 6-Oacetylnimbandiol, and nimbolide, replicate the binding mode of the Pin1 substrate peptide, interacting with high affinity with the binding site and thus destabilizing the Pin1 structure. Conclusion: Our results are supported by experimental data, and provide interesting structural insights into their molecular mechanism of action, indicating that their structural scaffolds could be used as a start point to develop new inhibitors against Pin1.

Calculation of free energy changes due to mutations from alchemical free energy simulations

2015 ◽  
Vol 14 (03) ◽  
pp. 1550023 ◽  
Author(s):  
M. Harunur Rashid ◽  
Germano Heinzelmann ◽  
Serdar Kuyucak
Keyword(s):  
Free Energy ◽  
Fundamental Problem ◽  
Binding Free Energy ◽  
Computational Design ◽  
Md Simulations ◽  
Binding Mode ◽  
Free Energy Calculations ◽  
Energy Calculations ◽  
Energy Simulations ◽  
Alchemical Free Energy

How a mutation affects the binding free energy of a ligand is a fundamental problem in molecular biology/biochemistry with many applications in pharmacology and biotechnology, e.g. design of drugs and enzymes. Free energy change due to a mutation can be determined most accurately by performing alchemical free energy calculations in molecular dynamics (MD) simulations. Here we discuss the necessary conditions for success of free energy calculations using toxin peptides that bind to ion channels as examples. We show that preservation of the binding mode is an essential requirement but this condition is not always satisfied, especially when the mutation involves a charged residue. Otherwise problems with accuracy of results encountered in mutation of charged residues can be overcome by performing the mutation on the ligand in the binding site and bulk simultaneously and in the same system. The proposed method will be useful in improving the affinity and selectivity profiles of drug leads and enzymes via computational design and protein engineering.

scholarly journals Binding mode and free energy prediction of fisetin/β-cyclodextrin inclusion complexes

2014 ◽  
Vol 10 ◽  
pp. 2789-2799 ◽  
Author(s):  
Bodee Nutho ◽  
Wasinee Khuntawee ◽  
Chompoonut Rungnim ◽  
Piamsook Pongsawasdi ◽  
Peter Wolschann ◽  
...  
Keyword(s):  
Free Energy ◽  
Molecular Docking ◽  
Phenyl Ring ◽  
Binding Free Energy ◽  
Md Simulations ◽  
Binding Mode ◽  
Free Energy Calculations ◽  
Energy Prediction ◽  
Theoretical Approaches ◽  
Preferential Binding

In the present study, our aim is to investigate the preferential binding mode and encapsulation of the flavonoid fisetin in the nano-pore of β-cyclodextrin (β-CD) at the molecular level using various theoretical approaches: molecular docking, molecular dynamics (MD) simulations and binding free energy calculations. The molecular docking suggested four possible fisetin orientations in the cavity through its chromone or phenyl ring with two different geometries of fisetin due to the rotatable bond between the two rings. From the multiple MD results, the phenyl ring of fisetin favours its inclusion into the β-CD cavity, whilst less binding or even unbinding preference was observed in the complexes where the larger chromone ring is located in the cavity. All MM- and QM-PBSA/GBSA free energy predictions supported the more stable fisetin/β-CD complex of the bound phenyl ring. Van der Waals interaction is the key force in forming the complexes. In addition, the quantum mechanics calculations with M06-2X/6-31G(d,p) clearly showed that both solvation effect and BSSE correction cannot be neglected for the energy determination of the chosen system.

scholarly journals Solvents to Fragments to Drugs: MD Applications in Drug Design

Molecules ◽  
2018 ◽  
Vol 23 (12) ◽  
pp. 3269 ◽  
Author(s):  
Lucas Defelipe ◽  
Juan Arcon ◽  
Carlos Modenutti ◽  
Marcelo Marti ◽  
Adrián Turjanski ◽  
...  
Keyword(s):  
Binding Free Energy ◽  
Mixed Solvents ◽  
Md Simulations ◽  
Binding Mode ◽  
Condensed State ◽  
Protein Targets ◽  
Drug Candidates ◽  
Interaction Sites ◽  
Aqueous Solvation ◽  
The Common

Simulations of molecular dynamics (MD) are playing an increasingly important role in structure-based drug discovery (SBDD). Here we review the use of MD for proteins in aqueous solvation, organic/aqueous mixed solvents (MDmix) and with small ligands, to the classic SBDD problems: Binding mode and binding free energy predictions. The simulation of proteins in their condensed state reveals solvent structures and preferential interaction sites (hot spots) on the protein surface. The information provided by water and its cosolvents can be used very effectively to understand protein ligand recognition and to improve the predictive capability of well-established methods such as molecular docking. The application of MD simulations to the study of the association of proteins with drug-like compounds is currently only possible for specific cases, as it remains computationally very expensive and labor intensive. MDmix simulations on the other hand, can be used systematically to address some of the common tasks in SBDD. With the advent of new tools and faster computers we expect to see an increase in the application of mixed solvent MD simulations to a plethora of protein targets to identify new drug candidates.

scholarly journals Sulfonanilide Derivatives in Identifying Novel Aromatase Inhibitors by Applying Docking, Virtual Screening, and MD Simulations Studies

2017 ◽  
Vol 2017 ◽  
pp. 1-17 ◽  
Author(s):  
Shailima Rampogu ◽  
Minky Son ◽  
Chanin Park ◽  
Hyong-Ha Kim ◽  
Jung-Keun Suh ◽  
...  
Keyword(s):  
Aromatase Inhibitors ◽  
Causes Of Death ◽  
Binding Free Energy ◽  
Chemical Compounds ◽  
Md Simulations ◽  
Free Energy Calculations ◽  
Lipinski’S Rule ◽  
Discovery Studio ◽  
Drug Candidates ◽  
Lipinski’S Rule Of Five

Breast cancer is one of the leading causes of death noticed in women across the world. Of late the most successful treatments rendered are the use of aromatase inhibitors (AIs). In the current study, a two-way approach for the identification of novel leads has been adapted. 81 chemical compounds were assessed to understand their potentiality against aromatase along with the four known drugs. Docking was performed employing the CDOCKER protocol available on the Discovery Studio (DS v4.5). Exemestane has displayed a higher dock score among the known drug candidates and is labeled as reference. Out of 81 ligands 14 have exhibited higher dock scores than the reference. In the second approach, these 14 compounds were utilized for the generation of the pharmacophore. The validated four-featured pharmacophore was then allowed to screen Chembridge database and the potential Hits were obtained after subjecting them to Lipinski’s rule of five and the ADMET properties. Subsequently, the acquired 3,050 Hits were escalated to molecular docking utilizing GOLD v5.0. Finally, the obtained Hits were consequently represented to be ideal lead candidates that were escalated to the MD simulations and binding free energy calculations. Additionally, the gene-disease association was performed to delineate the associated disease caused by CYP19A1.

Insights to the Binding of a Selective Adenosine A3 Receptor Antagonist using Molecular Dynamics Simulations, Binding Free Energy Calculations and Mutagenesis

2019 ◽  
Author(s):  
Panagiotis Lagarias ◽  
Kerry Barkan ◽  
Eva Tzortzini ◽  
Eleni Vrontaki ◽  
Margarita Stampelou ◽  
...  
Keyword(s):  
Free Energy ◽  
Computational Model ◽  
Molecular Mechanics ◽  
Surface Area ◽  
Binding Free Energy ◽  
Md Simulations ◽  
Free Energy Calculations ◽  
Binding Energies ◽  
Energy Calculations ◽  
Binding Free Energy Calculations

<p>Adenosine A<sub>3 </sub>receptor (A<sub>3</sub>R), is a promising drug target against cancer cell proliferation. Currently there is no experimentally determined structure of A<sub>3</sub>R. Here, we have investigate a computational model, previously applied successfully for agonists binding to A<sub>3</sub>R, using molecular dynamic (MD) simulations, Molecular Mechanics-Poisson Boltzmann Surface Area (MM-PBSA) and Molecular Mechanics-Generalized Born Surface Area (MM-GBSA) binding free energy calculations. Extensive computations were performed to explore the binding profile of O4-{[3-(2,6-dichlorophenyl)-5-methylisoxazol-4-yl]carbonyl}-2-methyl-1,3-thiazole-4-carbohydroximamide (K18) to A<sub>3</sub>R. K18 is a new specific and competitive antagonist at the orthosteric binding site of A<sub>3</sub>R, discovered using virtual screening and characterized pharmacologically in our previous studies. The most plausible binding conformation for the dichlorophenyl group of K18 inside the A<sub>3</sub>R is oriented towards trans-membrane helices (TM) 5 and 6, according to the MM-PBSA and MM-GBSA binding free energy calculations, and by the previous results obtained by mutating residues of TM5, TM6 to alanine which reduce antagonist potency. The results from 14 site-directed mutagenesis experiments were interpreted using MD simulations and MM-GBSA calculations which show that the relative binding free energies of the mutant A<sub>3</sub>R - K18 complexes compare to the WT A<sub>3</sub>R are in agreement with the effect of the mutations, i.e. the reduction, maintenance or increase of antagonist potency. We show that when the residues V169<sup>5.30</sup>, M177<sup>5.38</sup>, I249<sup>6.54</sup> involved in direct interactions with K18 are mutated to alanine, the mutant A<sub>3</sub>R - K18 complexes reduce potency, increase the RMSD value of K18 inside the binding area and the MM-GBSA binding free energy compared to the WT A<sub>3</sub>R complex. Our computational model shows that other mutant A<sub>3</sub>R complexes with K18, including directly interacting residues, i.e. F168<sup>5.29</sup>A, L246<sup>6.51</sup>A, N250<sup>6.55</sup>A complexes with K18 are not stable. In these complexes of A<sub>3</sub>R mutated in directly interacting residues one or more of the interactions between K18 and these residues are lost. In agreement with the experiments, the computations show that, M174<sup>5.35</sup> a residue which does not make direct interactions with K18 is critical for K18 binding. A striking results is that the mutation of residue V169<sup>5.30</sup> to glutamic acid maintained antagonistic potency. This effect is in agreement with the binding free energy calculations and it is suggested that is due to K18 re-orientation but also to the plasticity of A<sub>3</sub>R binding area. The mutation of direct interacting L90<sup>3.32</sup> in the low region and the non-directly interacting L264<sup>7.35</sup> to alanine in the middle region increases the antagonistic potency, suggesting that chemical modifications of K18 can be applied to augment antagonistic potency. The calculated binding energies Δ<i>G</i><sub>eff</sub> values of K18 against mutant A<sub>3</sub>Rs displayed very good correlation with experimental potencies (pA<sub>2</sub> values). These results further approve the computational model for the description of K18 binding with critical residues of the orthosteric binding area which can have implications for the design of more effective antagonists based on the structure of K18.</p>

scholarly journals Computational Exploration for Lead Compounds That Can Reverse the Nuclear Morphology in Progeria

2017 ◽  
Vol 2017 ◽  
pp. 1-15 ◽  
Author(s):  
Shailima Rampogu ◽  
Ayoung Baek ◽  
Minky Son ◽  
Amir Zeb ◽  
Chanin Park ◽  
...  
Keyword(s):  
Binding Free Energy ◽  
Genetic Disorder ◽  
Md Simulations ◽  
Free Energy Calculations ◽  
Premature Aging ◽  
Farnesyltransferase Inhibitors ◽  
Docking Simulations ◽  
Lipinski’S Rule ◽  
Computational Exploration ◽  
The Stability

Progeria is a rare genetic disorder characterized by premature aging that eventually leads to death and is noticed globally. Despite alarming conditions, this disease lacks effective medications; however, the farnesyltransferase inhibitors (FTIs) are a hope in the dark. Therefore, the objective of the present article is to identify new compounds from the databases employing pharmacophore based virtual screening. Utilizing nine training set compounds along with lonafarnib, a common feature pharmacophore was constructed consisting of four features. The validated Hypo1 was subsequently allowed to screen Maybridge, Chembridge, and Asinex databases to retrieve the novel lead candidates, which were then subjected to Lipinski’s rule of 5 and ADMET for drug-like assessment. The obtained 3,372 compounds were forwarded to docking simulations and were manually examined for the key interactions with the crucial residues. Two compounds that have demonstrated a higher dock score than the reference compounds and showed interactions with the crucial residues were subjected to MD simulations and binding free energy calculations to assess the stability of docked conformation and to investigate the binding interactions in detail. Furthermore, this study suggests that the Hits may be more effective against progeria and further the DFT studies were executed to understand their orbital energies.

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