Molecular Restrainer: Energy Minimization of NMR-Derived Structures

Molecular Restrainer is a SAMSON Extension that lets you quickly refine NMR-derived molecular structures - including protein-ligand complexes with non-standard residues - without the need for custom force-field files. It combines the Universal Force Field (UFF) with NOE-derived distance restraints to deliver publication-ready models for the Protein Data Bank (PDB) deposition or your next molecular-design project.

Key Benefits:

• Seamless postprocessing of CYANA outputs.
• No topology or parameter files needed thanks to UFF.
• Works with any molecule type: standard and non-standard residues, ligands.
• Batch processing: minimize entire NMR ensembles in one run.
• Delivers high-quality, energy-minimized structures ready for journals, PDB deposition, and downstream molecular-design workflows.
• Can later be easily adapted to other restraint formats beyond CYANA.

Reference

To learn about methods and the validation used in the Molecular Restrainer, please refer to the paper¹.

Note

This SAMSON Extension is a collaboration between the Orts Group (University of Vienna) and OneAngstrom, the provider of the SAMSON platform.

Requirements

• SAMSON 2024 or newer.
• FIRE optimizer module.

Quick-Start Tutorial

All you need is your NMR ensemble and the corresponding, NOE-derived, upper-distance-limit file (.upl) generated by CYANA.

• Open Molecular Restrainer: Home > Apps > Biology > Molecular Restrainer (or use Find everything...).
• Load your NMR structure into SAMSON. Multiple conformations will appear as a path (trajectory) plus one corresponding structure.
• Click Set inside Molecular Restrainer to select the structure and, optionally, a path you wish to minimize.
• Choose your .upl file under Restraints.
• Press Start. Done!

Detailed Setup

1. Set the Input Structure

Load in SAMSON an NMR-derived structure or ensemble that you would like to minimize.

Note

The input file can contain an ensemble (multiple conformations) - it will appear as a path (a trajectory) and a single structure. The Molecular Restrainer can perform a batch minimization of the whole ensemble.

Input Structure Must Be Complete

UFF, used by the Molecular Restrainer, expects the input structure to be valid, i.e., there should be no missing atoms (neither heavy atoms nor hydrogens), and no alternate locations. Usually, the NMR-derived structures should be OK, but they might be missing atoms in terminal residues (the N-terminal amine protons and C-terminal "OXT" oxygen) - for that the Molecular restrainer provides the Fix N- and C-termini option that checks and fixes the termini (only termini with standard amino acid residues would be fixed) in the initial structure and for all the conformations along the path with respect to the new positions. You can just keep this option checked at all time.

Need extra structure validation? See Protein Preparation & Validation. But note that adding hydrogens using SAMSON might change the names of hydrogens (if they were not named with the standard convention in the first place) which might influence the restraints loaded from an .upl file since they are done based on the atom names. Also, if you have a path, the adding of hydrogens will not add them along the path but only based on the positions of the initial structure and you will loose the hydrogens in the path.

Now, you need to set the structure in the Molecular Restrainer:

• If nothing is selected in the document, and you click Set, then it will choose the whole document as the input, i.e. all the structures and paths, if present.
• If you want to minimize only a specific structure (e.g., without a path), then select it in the Document view and click Set.

You should see a brief description of the specified input structure:

2. Load NOE Restraints

Accepted format: NOE-derived upper-distance-limit (.upl) files. Restraints are matched by residue ID and atom name - including pseudoatoms (see below).

UPL Format

For the format of the upper distance limit file (.upl) please see CYANA wiki - Distance restraint file.

Pseudoatoms

Molecular Restrainer builds pseudoatoms internally on-the-fly from hydrogen positions using these rules:

Pseudoatom	Parent atoms
Q	H1, H2, H3
QX	HX1, HX2, HX3, where X is a letter
QQX	HX11, HX12, HX13, HX21, HX22, HX23, where X is a letter

Partial sets (e.g., only HX2 and HX3 for a CH2 group) are accepted. Example: QA would be formed from HA1, HA2, HA3, if these are present. These rules correctly create most pseudoatoms of the standard residues.

Ligand Pseudoatoms

CYANA pseudoatom names for ligands and non-standard residues may not follow standard rules and might not be compatible with the abovementioned rules, hence, they might not be identified; if such restraints cannot be applied then they are reported in restraints.log.

Energy Minimization

Minimization parameters

To perform the energy minimization with restraints, the Molecular Restrainer uses a combined potential of the UFF and the restraints (see the paper¹) based on the specified weights:

Parameter	Default	Purpose
UFF weight	0.2	Balances generic force-field terms
Restraints weight	1.0	Keeps NOE distances satisfied
Random force weight	0.0	Helps escape local minima during the minimization (optional)

Leave defaults unless you have a specific need.

Stopping criteria

Minimization stops when either the restraint energy or the total energy plateaus - i.e., the relative energy change (\(\Delta E_{\text{rel}}\)) between successive iterations stays below to the given energy tolerance for a set number of iterations (plateau threshold).

You can leave the parameters set by default.

Relative Energy Change Formula

The relative energy change is computed as follows:

\[ \Delta E_{\text{rel}} = \frac{|E_{n+1} - E_n|}{|E_{n+1}|} \]

where \(E\) is either the restraints energy or the full energy and \(n\) is the integration step.

Tip

You can adjust plateau threshold or energy tolerance during the minimization if needed.

Run the minimization

• Pick an output folder - each run creates a timestamped sub-folder.
• Click Start. After a quick initialization, the energy minimization will start. The log window shows progress per conformation, and the progress bar shows the progress if there are multiple initial conformations.

Note

Need to abort? Click Stop; the current structure is still saved both in the results folder and as a conformation in the document.

The results folder will contain the following files per conformation:

• input.pdb - original system,
• energy.csv - table with energies vs iteration,
• minimized.pdb or stopped.pdb - final coordinates,

and the following global files:

• output.log - full console log,
• restraints.log - restraints that could not be applied because the atoms corresponding to a restraint where not found in the system,
• minimized_path.pdb - all minimized conformations as a single trajectory (if applicable).

Visualization of the restraints

During a run, Molecular Restrainer adds to the document a simulator with an interaction model that represent the combined potential of the UFF and the restraints. This model also visualizes the restraints - they are shown as additional "bonds" with their colors ranging from red (unsatisfied restraint, large energy) to green (satisfied restraint, small energy).

Want to modify how restraints are visualized (e.g., transparency, radius)? Selecting this interaction model in the Document view:

…then change settings in the Inspector.

Further Reading & Validation

For algorithm details, benchmarks, and validation, see the original paper¹.

References

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1. Jiří Mareš, Guneet Singh Tarang, Dmitriy Marin, Mehdi Mobli, Stephane Redon and Julien Orts. Streamlined Postprocessing of NMR Structures with the Molecular Restrainer: A Universal Tool for High-Quality Protein–Ligand Models and Non-Standard Amino Acid Residues. Int. J. Mol. Sci. 2025, 26(11), 5091. https://doi.org/10.3390/ijms26115091 ↩↩↩