Enhancing Hit Discovery in Virtual Screening through Absolute Protein–Ligand Binding Free-Energy Calculations

Abstract

In the hit identification stage of drug discovery, a diverse chemical space needs to be explored to identify initial hits. Contrary to empirical scoring functions, absolute protein−ligand binding free-energy perturbation (ABFEP) provides a theoretically more rigorous and accurate description of protein−ligand binding thermodynamics and could, in principle, greatly improve the hit rates in virtual screening. In this work, we describe an implementation of an accurate and reliable ABFEP method in FEP+. We validated the ABFEP method on eight congeneric compound series binding to eight protein receptors including both neutral and charged ligands. For ligands with net charges, the alchemical ion approach is adopted to avoid artifacts in electrostatic potential energy calculations. The calculated binding free energies correlate with experimental results with a weighted average of R 2 = 0.55 for the entire dataset. We also observe an overall root-mean-square error (RMSE) of 1.1 kcal/mol after shifting the zero-point of the simulation data to match the average experimental values. Through ABFEP calculations using apo versus holo protein structures, we demonstrated that the protein conformational and protonation state changes between the apo and holo proteins are the main physical factors contributing to the protein reorganization free energy manifested by the overestimation of raw ABFEP calculated binding free energies using the holo structures of the proteins. Furthermore, we performed ABFEP calculations in three virtual screening applications for hit enrichment. ABFEP greatly improves the hit rates as compared to docking scores or other methods like metadynamics. The good performance of ABFEP in rank ordering compounds demonstrated in this work confirms it as a useful tool to improve the hit rates in virtual screening, thus facilitating hit discovery.

Comparison of Receptor–Ligand Restraint Schemes for Alchemical Absolute Binding Free Energy Calculations

“…Algorithms have been proposed for the selection of Boresch restraints. ,,,− At a minimum, these aim to select stable restraints based on the geometry of the complex, while more sophisticated methods aim to enhance convergence by directly (e.g., based on H-bonds) or indirectly (based on minimum total variance of the distance, angles, and dihedrals) mimicking strong receptor–ligand interactions based on a short unrestrained simulation. However, there is no obviously superior method which has been shown to guarantee selection of numerically stable restraints with optimal convergence properties.…”

“…In general, it is nontrivial to select the optimum receptor–ligand restraints. Furthermore, ABFE calculations can be challenging to converge and therefore computationally costly because the ligand is completely removed. , As a result, application studies still combine RBFE and ABFE, with ABFE applied more successfully to low molecular-weight compounds . Thus, there are barriers to the routine application of ABFE calculations.…”

“…The performance and accessibility of ABFE calculations would be improved if it was trivial to select receptor–ligand restraints which resulted in stable simulations and produced optimal convergence. While progress has been made in this direction with tools for automated or partially automated restraint selection, − ,, there is still no restraint type or selection method which completely solves this issue.…”

Quantitatively Accounting for Protein Reorganization in Computer-Aided Drug Design

“…All simulations were performed in the μVT grand canonical ensemble, using grand canonical Monte Carlo (GCMC) with an excess chemical potential of −6.137 kcal/mol and a solvent number density of 0.03262 molecules/ Å 3 . 69 The AB-FEP 20 calculations used 68 and 108 replicas for neutral and charged ligands, respectively. The RB-FEP 14 calculations utilized 24 replicas and ran for 10 ns.…”

“…While absolute protein–ligand binding free-energy perturbation (AB-FEP) in principle can capture the apo-to-holo protein reorganization contribution to the ligand binding free energy, the long timescale associated with the conformational changes cannot be practically sampled in the relatively short AB-FEP simulations. This is why multiple studies have shown how sensitive AB-FEP results can be to different starting protein conformations. ,− These calculations are thus interpreted as absolute binding free energies to a specific protein conformation, and not the absolute binding free energy in vitro (Δ G exp ). The protein reorganization free energy from the apo-to-holo protein conformations is the missing piece.…”