A docking workflow can appear to fail for reasons that have little to do with the docking method or the ligand. The protein structure itself may contain alternate atom positions, missing hydrogens, unnecessary molecules, or geometry problems. These details are easy to overlook—and can make a structure harder to use reliably downstream.
Protein preparation is not simply a command to “clean everything.” The useful first step is deciding what your next workflow needs, then checking that the structure is suitable for it. In SAMSON, you can take a quick route for routine cleanup or inspect and repair specific issues when the structure needs more attention.
A practical first pass
Before changing a structure, keep an untouched copy of the source file. Then ask what should remain for your particular task. A water molecule, ion, cofactor, or ligand may be unnecessary for one calculation but important in another. This matters because the one-click preparation tool can delete ligands—including covalently attached ligands—cofactors, and other small molecules, as well as strip waters and clear monatomic ions.
For a structure that is already reasonably complete, use Home > Prepare. Its options address common housekeeping steps:
- Alternate locations: retains the highest-occupancy atoms. When occupancies are tied or absent, it prefers alternate location A if present, then B, and so on.
- Unneeded molecules: removes selected ligands and other small molecules, waters, and monatomic ions.
- Hydrogens: adds hydrogens according to residue type for standard residues, or according to valence for other atoms.
The key is to review which options suit your system rather than treating every removal as universally appropriate.
When a quick cleanup is not enough
A structure can look tidy and still have issues worth checking. Open Home > Validate to inspect alternate locations, bond lengths, non-standard residues, and clashes or contacts. This helps separate simple cleanup from problems that may need a more targeted fix.
For example, alternate locations can be reviewed in the Alt. locations tab: find them, inspect the list, and remove them when appropriate. Bond-length checks can reveal bonds outside expected ranges. The module can also identify non-standard residue aliases and highlight steric clashes. If side-chain clashes need attention, the documentation points to the Rotamers editor, which changes side chains using a backbone-dependent rotamer library.
Validation is also a useful moment to consider the downstream workflow. A docking setup may have different requirements from a molecular-dynamics system. For instance, the documentation notes that AutoDock Vina needs only polar hydrogens for determining hydrogen bonds, so minimizing a structure after adding hydrogens is not necessarily required for every task.
Missing atoms or residues call for a deeper fix
If residues or atoms are missing, routine preparation is not the same as rebuilding them. The PDBFixer extension can add missing residues based on SEQRES records, add missing heavy atoms and side chains, convert non-standard residues to standard equivalents, and add hydrogens at a specified pH. It can also remove water or heterogens, resolve alternate locations, and build explicit-solvent water boxes or lipid membranes.
A simple decision rule helps: use Prepare for routine cleanup of a structure that is already suitable; use Validate when you need to identify or inspect specific issues; and consider PDBFixer when atoms or residues need to be built or protonation needs to be set for a particular pH. For many structures, that sequence makes preparation feel less like guesswork and more like a short, deliberate checklist.
For the complete workflow and detailed options, see the SAMSON protein preparation and validation tutorial.
Note: SAMSON and all SAMSON Extensions are free for non-commercial use. Get SAMSON.