Predicted Binding Free Energies Research Articles

Accelerated Enveloping Distribution Sampling (AEDS) Allows for Efficient Sampling of Orthogonal Degrees of Freedom.

One of the most challenging aspects in the molecular simulation of proteins is the study of slowly relaxing processes, such as loop rearrangements or ligands that adopt different conformations in the binding site. State-of-the-art methods used to calculate binding free energies rely on performing several short simulations (lambda steps), in which the ligand is slowly transformed into the endstates of interest. This makes capturing the slowly relaxing processes even more difficult, as they would need to be observed in all of the lambda steps. One attractive alternative is the use of a reference state capable of sampling all of the endstates of interest in a single simulation. However, the energy barriers between the states can be too high to overcome, thus hindering the sampling of all of the relevant conformations. Accelerated enveloping distribution sampling (AEDS) is a recently developed reference state technique that circumvents the high-energy-barrier challenge by adding a boosting potential that flattens the energy landscape without distorting the energy minima. In the present work, we apply AEDS to the well-studied benchmark system T4 lysozyme L99A. The T4 lysozyme L99A mutant contains a hydrophobic pocket in which there is a valine (valine 111), whose conformation influences the binding efficiencies of the different ligands. Incorrectly sampling the dihedral angle can lead to errors in predicted binding free energies of up to 16 kJ mol-1. This protein constitutes an ideal scenario to showcase the advantages and challenges when using AEDS in the presence of slow relaxing processes. We show that AEDS is able to successfully sample the relevant degrees of freedom, providing accurate binding free energies, without the need of previous information of the system in the form of collective variables. Additionally, we showcase the capabilities of AEDS to efficiently screen a set of ligands. These results represent a promising first step toward the development of free-energy methods that can respond to more intricate molecular events.

AMBER Drug Discovery Boost Tools: Automated Workflow for Production Free-Energy Simulation Setup and Analysis (ProFESSA).

We report an automated workflow for production free-energy simulation setup and analysis (ProFESSA) using the GPU-accelerated AMBER free-energy engine with enhanced sampling features and analysis tools, part of the AMBER Drug Discovery Boost package that has been integrated into the AMBER22 release. The workflow establishes a flexible, end-to-end pipeline for performing alchemical free-energy simulations that brings to bear technologies, including new enhanced sampling features and analysis tools, to practical drug discovery problems. ProFESSA provides the user with top-level control of large sets of free-energy calculations and offers access to the following key functionalities: (1) automated setup of file infrastructure; (2) enhanced conformational and alchemical sampling with the ACES method; and (3) network-wide free-energy analysis with the optional imposition of cycle closure and experimental constraints. The workflow is applied to perform absolute and relative solvation free-energy and relative ligand-protein binding free-energy calculations using different atom-mapping procedures. Results demonstrate that the workflow is internally consistent and highly robust. Further, the application of a new network-wide Lagrange multiplier constraint analysis that imposes key experimental constraints substantially improves binding free-energy predictions.

Evaluating the Reliability of MM-PB/GB-SA Method for the Protein-Ligand Binding Free Energies Using Penicillopepsin-Inhibitor ligands

An accurate prediction of the ligand-receptor binding free energies (ΔG) is a critical step in the early stages of rational drug design. The Molecular Mechanics-Generalized Born Surface Area (MM-GBSA) method is a popular
 
 approach to estimate ΔG. However, correlations between the predicted and the experimental ΔG are variable. The goal of this study is to investigate various approaches to optimize accuracy of the MM-GBSA method. A molecular dynamic (MD) simulations protocol was applied using penicillopepsin receptor against its inhibitor ligands, repeated 50 times for each complex system. After that, ΔG of the five inhibitors were predicted using MM-GBSA method. Moreover, a diverse ΔG values were calculated from the replicate MD simulations of each system. The results were showed correlations not only between the predicted and the experimental binding affinities of the systems but also between the predicted values and root-mean-square deviation. In addition, statistical analysis was evaluated the sample size.

Hierarchical Virtual Screening and Binding Free Energy Prediction of Potential Modulators of Aedes Aegypti Odorant-Binding Protein 1.

The Aedes aegypti mosquito is the main hematophagous vector responsible for arbovirus transmission in Brazil. The disruption of A. aegypti hematophagy remains one of the most efficient and least toxic methods against these diseases and, therefore, efforts in the research of new chemical entities with repellent activity have advanced due to the elucidation of the functionality of the olfactory receptors and the behavior of mosquitoes. With the growing interest of the pharmaceutical and cosmetic industries in the development of chemical entities with repellent activity, computational studies (e.g., virtual screening and molecular modeling) are a way to prioritize potential modulators with stereoelectronic characteristics (e.g., pharmacophore models) and binding affinity to the AaegOBP1 binding site (e.g., molecular docking) at a lower computational cost. Thus, pharmacophore- and docking-based virtual screening was employed to prioritize compounds from Sigma-Aldrich® (n = 126,851) and biogenic databases (n = 8766). In addition, molecular dynamics (MD) was performed to prioritize the most potential potent compounds compared to DEET according to free binding energy calculations. Two compounds showed adequate stereoelectronic requirements (QFIT > 81.53), AaegOBP1 binding site score (Score > 42.0), volatility and non-toxic properties and better binding free energy value (∆G < −24.13 kcal/mol) compared to DEET ((N,N-diethyl-meta-toluamide)) (∆G = −24.13 kcal/mol).

Absolute Binding Free Energy Calculations for Buried Water Molecules.

Water often plays a key role in mediating protein-ligand interactions. Understanding contributions from active-site water molecules to binding thermodynamics of a ligand is important in predicting binding free energies for ligand optimization. In this work, we tested a non-equilibrium switching method for absolute binding free energy calculations on water molecules in binding sites of 13 systems. We discuss the lessons we learned about identified issues that affected our calculations and ways to address them. This work fits with our larger focus on how to do accurate ligand binding free energy calculations when water rearrangements are very slow, such as rearrangements due to ligand modification (as in relative free energy calculations) or ligand binding (as in absolute free energy calculations). The method studied in this work can potentially be used to account for limited water sampling via providing endpoint corrections to free energy calculations using our calculated binding free energy of water.

The impact of simulation time in predicting binding free energies using end-point approaches.

The profound impact of <i>in silico</i> studies for a fast-paced drug discovery pipeline is undeniable for pharmaceutical community. The rational design of novel drug candidates necessitates considering optimization of their different aspects prior to synthesis and biological evaluations. The affinity prediction of small ligands to target of interest for rank-ordering the potential ligands is one of the most routinely used steps in the context of virtual screening. So, the end-point methods were employed for binding free energy estimation focusing on evaluating simulation time effect. Then, a set of human aldose reductase inhibitors were selected for molecular dynamics (MD)-based binding free energy calculations. A total of 100[Formula: see text]ns MD simulation time was conducted for the ligand-receptor complexes followed by prediction of binding free energies using MM/PB(GB)SA and LIE approaches under different simulation time. The results revealed that a maximum of 30[Formula: see text]ns simulation time is sufficient for determination of binding affinities inferred from steady trend of squared correlation values (R²) between experimental and predicted [Formula: see text]G as a function of MD simulation time. In conclusion, the MM/PB(GB)SA algorithms performed well in terms of binding affinity prediction compared to LIE approach. The results provide new insights for large-scale applications of such predictions in an affordable computational cost.

An overview of the SAMPL8 host–guest binding challenge

The SAMPL series of challenges aim to focus the community on specific modeling challenges, while testing and hopefully driving progress of computational methods to help guide pharmaceutical drug discovery. In this study, we report on the results of the SAMPL8 host–guest blind challenge for predicting absolute binding affinities. SAMPL8 focused on two host–guest datasets, one involving the cucurbituril CB8 (with a series of common drugs of abuse) and another involving two different Gibb deep-cavity cavitands. The latter dataset involved a previously featured deep cavity cavitand (TEMOA) as well as a new variant (TEETOA), both binding to a series of relatively rigid fragment-like guests. Challenge participants employed a reasonably wide variety of methods, though many of these were based on molecular simulations, and predictive accuracy was mixed. As in some previous SAMPL iterations (SAMPL6 and SAMPL7), we found that one approach to achieve greater accuracy was to apply empirical corrections to the binding free energy predictions, taking advantage of prior data on binding to these hosts. Another approach which performed well was a hybrid MD-based approach with reweighting to a force matched QM potential. In the cavitand challenge, an alchemical method using the AMOEBA-polarizable force field achieved the best success with RMSE less than 1 kcal/mol, while another alchemical approach (ATM/GAFF2-AM1BCC/TIP3P/HREM) had RMSE less than 1.75 kcal/mol. The work discussed here also highlights several important lessons; for example, retrospective studies of reference calculations demonstrate the sensitivity of predicted binding free energies to ethyl group sampling and/or guest starting pose, providing guidance to help improve future studies on these systems.

Exploring the Binding Affinity and Mechanism between ACE2 and the Trimers of Delta and Omicron Spike Proteins by Molecular Dynamics Simulation and Bioassay.

Five major variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have emerged and posed challenges in controlling the pandemic. Among them, the current dominant variant, viz., Omicron, has raised serious concerns about its infectiousness and antibody neutralization. However, few studies pay attention to the effect of the mutations on the dynamic interaction network of Omicron S protein trimers binding to the host angiotensin-converting enzyme 2 (ACE2). In this study, we conducted molecular dynamics (MD) simulations and enzyme linked immunosorbent assay (ELISA) to explore the binding strength and mechanism of wild type (WT), Delta, and Omicron S protein trimers to ACE2. The results showed that the binding capacities of both the two variants' S protein trimers to ACE2 are enhanced in varying degrees, indicating possibly higher cell infectiousness. Energy decomposition and protein-protein interaction network analysis suggested that both the mutational and conserved sites make effects on the increase in the overall affinity through a variety of interactions. The experimentally determined KD values by biolayer interferometry (BLI) and the predicted binding free energies of the RBDs of Delta and Omicron to mAb HLX70 revealed that the two variants may have the high risk of immune evasion from the mAb. These results are not only helpful in understanding the binding strength and mechanism of S protein trimer-ACE2 but also beneficial for drug, especially for antibody development.

3D-RISM-AI: A Machine Learning Approach to Predict Protein-Ligand Binding Affinity Using 3D-RISM.

Hydration free energy (HFE) is a key factor in improving protein–ligand binding free energy (BFE) prediction accuracy. The HFE itself can be calculated using the three-dimensional reference interaction model (3D-RISM); however, the BFE predictions solely evaluated using 3D-RISM are not correlated to the experimental BFE for abundant protein–ligand pairs. In this study, to predict the BFE for multiple sets of protein–ligand pairs, we propose a machine learning approach incorporating the HFEs obtained using 3D-RISM, termed 3D-RISM-AI. In the learning process, structural metrics, intra-/intermolecular energies, and HFEs obtained via 3D-RISM of ∼4000 complexes in the PDBbind database (ver. 2018) were used. The BFEs predicted using 3D-RISM-AI were well correlated to the experimental data (Pearson’s correlation coefficient of 0.80 and root-mean-square error of 1.91 kcal/mol). As important factors for the prediction, the difference in the solvent accessible surface area between the bound and unbound structures and the hydration properties of the ligands were detected during the learning process.

Nonbonded Force Field Parameters from Minimal Basis Iterative Stockholder Partitioning of the Molecular Electron Density Improve CB7 Host-Guest Affinity Predictions.

Binding affinity prediction by means of computer simulation has been increasingly incorporated in drug discovery projects. Its wide application, however, is limited by the prediction accuracy of the free energy calculations. The main error sources are force fields used to describe molecular interactions and incomplete sampling of the configurational space. Organic host-guest systems have been used to address force field quality because they share similar interactions found in ligands and receptors, and their rigidity facilitates configurational sampling. Here, we test the binding free energy prediction accuracy for 14 guests with an aromatic or adamantane core and the CB7 host using molecular electron density derived nonbonded force field parameters. We developed a computational workflow written in Python to derive atomic charges and Lennard-Jones parameters with the Minimal Basis Iterative Stockholder method using the polarized electron density of several configurations of each guest in the bound and unbound states. The resulting nonbonded force field parameters improve binding affinity prediction, especially for guests with an adamantane core in which repulsive exchange and dispersion interactions to the host dominate.

Understanding gilteritinib resistance to FLT3-F691L mutation through an integrated computational strategy.

FMS-like tyrosine kinase 3 (FLT3) serves as an important drug target for acute myeloid leukemia (AML), and gene mutations of FLT3 have been closely associated with AML patients with an incidence rate of ~ 30%. However, the mechanism of the clinically relevant F691L gatekeeper mutation conferred resistance to the drug gilteritinib remained poorly understood. In this study, multiple microsecond molecular dynamics (MD) simulations, end-point free energy calculations, and dynamic correlated and network analyses were performed to investigate the molecular basis of gilteritinib resistance to the FLT3-F691L mutation. The simulations revealed that the resistant mutation largely induced the conformational changes of the activation loop (A-loop), the phosphate-binding loop, and the helix αC of the FLT3 protein. The binding abilities of the gilteritinib to the wild-type and the F691L mutant were different through the binding free energy prediction. The simulation results further indicated that the driving force to determine the binding affinity of gilteritinib was derived from the differences in the energy terms of electrostatic and van der Waals interactions. Moreover, the per-residue free energy decomposition suggested that the four residues (Phe803, Gly831, Leu832, and Ala833) located at the A-loop of FLT3 had a significant impact on the binding affinity of gilteritinib to the F691L mutant. This study may provide useful information for the design of novel FLT3 inhibitors specially targeting the F691L gatekeeper mutant.

Mechanistic Insights into the Mechanism of Inhibitor Selectivity toward the Dark Kinase STK17B against Its High Homology STK17A.

As a member of the death-associated protein kinase (DAPK) family, STK17B plays an important role in the regulation of cellular apoptosis and has been considered as a promising drug target for hepatocellular carcinoma. However, the highly conserved ATP-binding site of protein kinases represents a challenge to design selective inhibitors for a specific DAPK isoform. In this study, molecular docking, multiple large-scale molecular dynamics (MD) simulations, and binding free energy calculations were performed to decipher the molecular mechanism of the binding selectivity of PKIS43 toward STK17B against its high homology STK17A. MD simulations revealed that STK17A underwent a significant conformational arrangement of the activation loop compared to STK17B. The binding free energy predictions suggested that the driving force to control the binding selectivity of PKIS43 was derived from the difference in the protein–ligand electrostatic interactions. Furthermore, the per-residue free energy decomposition unveiled that the energy contribution from Arg41 at the phosphate-binding loop of STK17B was the determinant factor responsible for the binding specificity of PKIS43. This study may provide useful information for the rational design of novel and potent selective inhibitors toward STK17B.

Automation of AMOEBA polarizable force field for small molecules: Poltype 2.

A next-generation protocol (Poltype 2) has been developed which automatically generates AMOEBA polarizable force field parameters for small molecules. Both features and computational efficiency have been drastically improved. Notable advances include improved database transferability using SMILES, robust torsion fitting, non-aromatic ring torsion parameterization, coupled torsion-torsion parameterization, Van der Waals parameter refinement using ab initio dimer data and an intelligent fragmentation scheme that produces parameters with dramatically reduced ab initio computational cost. Additional improvements include better local frame assignment for atomic multipoles, automated formal charge assignment, Zwitterion detection, smart memory resource defaults, parallelized fragment job submission, incorporation of Psi4 quantum package, ab initio error handling, ionization state enumeration, hydration free energy prediction and binding free energy prediction. For validation, we have applied Poltype 2 to ~1000 FDA approved drug molecules from DrugBank. The ab initio molecular dipole moments and electrostatic potential values were compared with Poltype 2 derived AMOEBA counterparts. Parameters were further substantiated by calculating hydration free energy (HFE) on 40 small organic molecules and were compared with experimental data, resulting in an RMSE error of 0.59 kcal/mol. The torsion database has expanded to include 3543 fragments derived from FDA approved drugs. Poltype 2 provides a convenient utility for applications including binding free energy prediction for computational drug discovery. Further improvement will focus on automated parameter refinement by experimental liquid properties, expansion of the Van der Waals parameter database and automated parametrization of modified bio-fragments such as amino and nucleic acids.

The performance of ensemble-based free energy protocols in computing binding affinities to ROS1 kinase

Optimization of binding affinities for compounds to their target protein is a primary objective in drug discovery. Herein we report on a collaborative study that evaluates a set of compounds binding to ROS1 kinase. We use ESMACS (enhanced sampling of molecular dynamics with approximation of continuum solvent) and TIES (thermodynamic integration with enhanced sampling) protocols to rank the binding free energies. The predicted binding free energies from ESMACS simulations show good correlations with experimental data for subsets of the compounds. Consistent binding free energy differences are generated for TIES and ESMACS. Although an unexplained overestimation exists, we obtain excellent statistical rankings across the set of compounds from the TIES protocol, with a Pearson correlation coefficient of 0.90 between calculated and experimental activities.

In-silico discovery of inhibitors against human papillomavirus E1 protein

High-risk (HR) Human papillomavirus (e.g. HPV16 and HPV18) causes approximately two-thirds of all cervical cancers in women. Although the first and second-generation vaccines confer some protection against individuals, there are no approved drugs to treat HR-HPV infections to-date. The HPV E1 protein is an attractive drug target because the protein is highly conserved across all HPV types and is crucial for the regulation of viral DNA replication. Hence, we used the Random Forest algorithm to construct a Quantitative-Structure Activity Relationship (QSAR) model to predict the potential inhibitors against the HPV E1 protein. Our QSAR classification model achieved an accuracy of 87.5%, area under the receiver operating characteristic curve of 1.00, and F-measure of 0.87 when evaluated using an external test set. We conducted a drug repurposing campaign by deploying the model to screen the Drugbank database. The top three compounds, namely Cinalukast, Lobeglitazone, and Efatutazone were analyzed for their cell membrane permeability, toxicity, and carcinogenicity. Finally, these three compounds were subjected to molecular docking and 200 ns-long Molecular Dynamics (MD) simulations. The predicted binding free energies for the candidates were calculated using the MM-GBSA method. The binding free energies for Cinalukast, Lobeglitazone, and Efatutazone were −37.84 kcal/mol, −25.30 kcal/mol, and −29.89 kcal/mol respectively. Therefore, we propose their chemical scaffolds for future rational design of E1 inhibitors. Communicated by Ramaswamy H. Sarma

Scoring Functions for Protein-Ligand Binding Affinity Prediction using Structure-Based Deep Learning: A Review.

The rapid and accurate in silico prediction of protein-ligand binding free energies or binding affinities has the potential to transform drug discovery. In recent years, there has been a rapid growth of interest in deep learning methods for the prediction of protein-ligand binding affinities based on the structural information of protein-ligand complexes. These structure-based scoring functions often obtain better results than classical scoring functions when applied within their applicability domain. Here we review structure-based scoring functions for binding affinity prediction based on deep learning, focussing on different types of architectures, featurization strategies, data sets, methods for training and evaluation, and the role of explainable artificial intelligence in building useful models for real drug-discovery applications.

Computational Molecular Modeling Studies of Some Mycobacterium Tuberculosis Alanine Racemase Inhibitors.

Alanine racemase is a pyridoxal-5'-phosphate dependent bacterial enzyme that provides the essential peptidoglycan precursor D-alanine, utilized for cell wall synthesis. This enzyme is ubiquitous throughout bacteria, including Mycobacterium tuberculosis, making it an attractive target for antibacterial drug discovery. We investigated the binding mode of twenty five reported Mycobacterium tuberculosis alanine racemase inhibitors. The results obtained from molecular docking studies emphasized the importance of inhibitor interaction with Lys42, Tyr46, Arg140, His172 and Tyr175 residues at the catalytic binding pocket of alanine racemase enzyme. The predicted binding free energies showed that van der Waals and nonpolar solvation interactions are the driving force for binding of inhibitors. Molecular dynamics simulation studies of four such inhibitor-alanine racemase systems were further explored to study the inhibition mechanism. The quantum chemical parameters calculated at the B3LYP/6-31G**++ level of theory indicated that the inhibitors must have low values of the lowest unoccupied molecular orbital energy and high values of electrostatic potential for stronger interactions. We expect that this study can provide significant theoretical guidance for design of potent Mycobacterium tuberculosis alanine racemase inhibitors in future.

Predicting Binding Free Energies for DPS Protein-DNA Complexes and Crystals Using Molecular Dynamics

The interaction between deoxyribonucleic acid (DNA) and deoxyribonucleic acid-binding protein from starved cells (DPS) in bacterial cells leads to intracellular crystallization of the genetic material of bacteria, which contributes to the survival of bacteria under stress factors, including antibacterial agents. Molecular modeling can help explain the molecular mechanisms of DNA binding to this protein. In this paper, we report a supercomputer simulation of the molecular dynamics of several types DNA-DPS complexes and crystals ranging from DPS+DNA dimer to DNA in periodic crystal channels of Escherichia coli DPS protein using a coarse-grained Martini force field. By modeling DNA of 24 base pairs, comparable in size to the diameter of the DPS protein, we use the slow-growth thermodynamic integration method to find binding protein-DNA free energy and discuss the contribution of ions and the length of trajectories sufficient for this type of simulations. The results obtained are important for further research in the field of simulation of biological DNA-protein crystals and the study of the molecular mechanisms of DNA interaction with the DPS protein.

Fast and Effective Prediction of the Absolute Binding Free Energies of Covalent Inhibitors of SARS-CoV-2 Main Protease and 20S Proteasome.

The COVID-19 pandemic has been a public health emergency with continuously evolving deadly variants around the globe. Among many preventive and therapeutic strategies, the design of covalent inhibitors targeting the main protease (Mpro) of SARS-CoV-2 that causes COVID-19 has been one of the hotly pursued areas. Currently, about 30% of marketed drugs that target enzymes are covalent inhibitors. Such inhibitors have been shown in recent years to have many advantages that counteract past reservation of their potential off-target activities, which can be minimized by modulation of the electrophilic warhead and simultaneous optimization of nearby noncovalent interactions. This process can be greatly accelerated by exploration of binding affinities using computational models, which are not well-established yet due to the requirement of capturing the chemical nature of covalent bond formation. Here, we present a robust computational method for effective prediction of absolute binding free energies (ABFEs) of covalent inhibitors. This is done by integrating the protein dipoles Langevin dipoles method (in the PDLD/S-LRA/β version) with quantum mechanical calculations of the energetics of the reaction of the warhead and its amino acid target, in water. This approach evaluates the combined effects of the covalent and noncovalent contributions. The applicability of the method is illustrated by predicting the ABFEs of covalent inhibitors of SARS-CoV-2 Mpro and the 20S proteasome. Our results are found to be reliable in predicting ABFEs for cases where the warheads are significantly different. This computational protocol might be a powerful tool for designing effective covalent inhibitors especially for SARS-CoV-2 Mpro and for targeted protein degradation.

Protein-Protein Binding Free Energy Predictions with the MM/PBSA Approach Complemented with the Gaussian-Based Method for Entropy Estimation.

Here, we present a Gaussian-based method for estimation of protein–protein binding entropy to augment the molecular mechanics Poisson–Boltzmann surface area (MM/PBSA) method for computational prediction of binding free energy (ΔG). The method is termed f5-MM/PBSA/E, where “E” stands for entropy and f5 for five adjustable parameters. The enthalpy components of ΔG (molecular mechanics, polar and non-polar solvation energies) are computed from a single implicit solvent generalized Born (GB) energy minimized structure of a protein–protein complex, while the binding entropy is computed using independently GB energy minimized unbound and bound structures. It should be emphasized that the f5-MM/PBSA/E method does not use snapshots, just energy minimized structures, and is thus very fast and computationally efficient. The method is trained and benchmarked in 5-fold validation test over a data set consisting of 46 protein–protein binding cases with experimentally determined dissociation constant Kd values. This data set has been used for benchmarking in recently published protein–protein binding studies that apply conventional MM/PBSA and MM/PBSA with an enhanced sampling method. The f5-MM/PBSA/E tested on the same data set achieves similar or better performance than these computationally demanding approaches, making it an excellent choice for high throughput protein–protein binding affinity prediction studies.