Absolute Binding Free Energy

Absolute Binding Free Energy (ABFE) calculates how strongly one ligand binds to a protein in water, reported as a binding free energy, ΔG_bind, in kcal/mol.

A more negative ΔG_bind means tighter binding. Because the value is absolute, not measured relative to another molecule, you can compare binders with completely different chemical scaffolds.

Under the hood it runs an alchemical free energy simulation: the ligand is gradually switched off from its surroundings both in plain water and inside the protein pocket, and the difference between those two gives the binding free energy.

When to use it

Use it when you want the binding strength of a single ligand to a target, on an absolute scale, or when you want to compare structurally diverse molecules where Relative Binding Free Energy, which needs closely related pairs, does not fit.

Inputs

Input	Required	What it is
`pdb_file`	yes	PDB file of the protein. Upload one, or fetch from RCSB by ID. It may already contain the ligand.
`ligand_smiles`	yes	SMILES string of the ligand.
`ligand_file`	no	A ligand structure file in SDF, PDB, or CIF format, if you have one.
`ligand_in_pdb_file`	no, default `false`	Set to `true` if the ligand is already inside the protein PDB (a cocrystal structure).
`ligand_chain_id`	no	Chain ID of the ligand in the PDB. Only needed when pulling the ligand out of a cocrystal PDB. Detected automatically if left blank.
`ligand_residue_name`	no	Residue name of the ligand in the PDB. Detected automatically if left blank.
`assign_protonation_states`	no, default `true`	Protonates the ligand automatically at the given pH. Turn off if your input is already protonated.
`ph`	no, default `7.0`	pH used to decide protonation.
`solvent_equil_length`, `solvent_prod_length`	no, default `0.02`, `0.1` ns	Equilibration and production length per replica, in plain solvent.
`complex_equil_length`, `complex_prod_length`	no, default `0.02`, `0.1` ns	Equilibration and production length per replica, for the ligand and protein complex.
`protocol_repeats`	no, default `3`	Number of independent repeats. More repeats give a better error estimate.
`platform`	no, default `CUDA`	Compute platform, one of `CUDA`, `OpenCL`, `CPU`, or `Reference`.
`keep_dirs`	no, default `true`	Preserves the full simulation outputs so you can download them. Turn off only if you do not need the raw data.

Longer simulation lengths and more repeats give more reliable numbers, but cost more runtime and credits.

How to run it

Submit your own protein and ligand from Azulene Studio, the Python SDK, or the CLI. New here? The Get started page walks through installing, logging in, and running a ready made example first.

In Azulene Studio

Open Absolute Binding Free Energy from the tools list, then on the Inputs and Parameters step upload your protein PDB (or fetch it from RCSB by ID), enter the ligand SMILES (or upload a ligand file), adjust the pH and simulation lengths if you want, then Review and Submit.

From the Python SDK

from opal import jobs

result = jobs.submit(
    job_type="absolute_binding",
    input_data={
        "pdb_file": "/path/to/your/protein.pdb",
        "ligand_smiles": "CC(=O)Oc1ccccc1C(=O)O",
        "ph": 7.0,
    },
)

From the CLI

Pass the inputs as a JSON string.

opal jobs submit --job-type absolute_binding \
  --input-data '{"pdb_file": "/path/to/your/protein.pdb", "ligand_smiles": "CC(=O)Oc1ccccc1C(=O)O", "ph": 7.0}'

Reading the result

The main output is dg_binding, the binding free energy ΔG_bind, in kcal/mol.

A more negative number means a stronger predicted binder. Rank a set of ligands by their dg_binding to find the tightest binders. If you keep the full simulation outputs, the raw per replica data is available to download as well.

Notes

Keep the simulation lengths short for a quick first run. For reliable results, use longer production lengths and at least 3 repeats. ABFE runs on a GPU, and runtime grows with the simulation lengths and the number of repeats.