BFEE-Docking: A User-Friendly and Customizable End-to-End Tool from High-Throughput Virtual Screening to Binding Free-Energy Calculations

Rational drug design typically relies on a sequential workflow comprising virtual screening, flexible redocking, and free energy calculations. Specialized tools exist for individual steps, including our Binding Free-Energy Estimator 3 (BFEE3) for absolute binding free-energy (ABFE) calculations. However, the field lacks an open-source platform that seamlessly connects virtual screening to rigorous free-energy evaluation while allowing nonspecialists to customize intermediate stages. Although commercial suites offer such integration, they often suffer from limited accessibility. To address this limitation, we develop BFEE-docking as a companion platform to BFEE3, extending BFEE3 toward an integrated, automated drug-discovery workflow. Requiring only a standard protein PDB file and a ligand database (SMI/SDF), this cross-platform software automates GPU-accelerated virtual screening, flexible redocking, and free-energy estimation using either MM-GBSA or BFEE3-based ABFE calculations. The software autonomously handles complex preparatory tasks, including pH-based protonation, chain cleaning, binding site identification, and file format conversion. Notably, BFEE-docking interfaces directly with PyMOL to provide real-time visualization of intermediate results, including binding site locations, docked ligand conformations, and flexible residue configurations. This feature allows the end-user, regardless of their computational background, to inspect and customize the protocol on the fly. We demonstrate the usefulness of BFEE-docking through a challenging inhibitor screening campaign against the epidermal growth factor receptor triple mutant (EGFR TM ), achieving formally exact binding free-energy evaluation within the screening workflow while preserving protonation-state-specific docking poses for separate MM-GBSA or ABFE calculations. The pipeline successfully identifies two novel potential inhibitors overlooked in our previous studies, showing the reliability and effectiveness of BFEE-docking in drug discovery.