Calculate RMSD between protein structures with automatic alignment Learn more
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Output
Configure inputs to begin
Set options on the left, then click “Calculate”.
Calculate RMSD between protein structures with automatic alignment Learn more
Configure inputs to begin
Set options on the left, then click “Calculate”.
Calculate RMSD between protein structures with automatic alignment Learn more
Configure inputs to begin
Set options on the left, then click “Calculate”.
Upload or fetch a reference structure and one or more comparison structures, choose the atoms to measure, then select Calculate. ProteinIQ matches corresponding coordinates, performs an optimal rigid-body fit, and returns global RMSD, matched coverage, per-residue RMSD, fitted structures, and the rotation and translation applied to each comparison.
For a verified example, fetch 1UBQ as the reference and 1D3Z as the comparison. Keep the default settings:
RMSD atom selection: Alpha carbons (CA)
Alignment atom selection: Same as RMSD selection
Compare by: Structure
RMSD weighting: Uniform
Multi-model structures: First model only
Automatic alignment: OnThe current RCSB coordinate files produce:
Reference: 1UBQ
Comparison: 1D3Z:model_1
RMSD: 0.521 Å
Measured atoms: 76
Fit atoms: 76
Reference coverage: 100%
Comparison coverage: 100%1D3Z contains 10 NMR models, so first-model mode also reports that only model 1 was used. Choose All model pairs to compare every 1D3Z model.
The 0.521 Å result is reproducible for the current RCSB PDBx/mmCIF files and the settings shown above. It compares all 76 matched Cα atoms in 1UBQ with model 1 of 1D3Z; it is not a general threshold for deciding whether two unrelated proteins have the same fold.
| Input | Accepted formats | Limit | Matching role |
|---|---|---|---|
| Reference structure | .pdb, .ent, .cif, .mmcif, .pdbx | One file, up to 50 MB | Fixed coordinate set |
| Comparison structures | .pdb, .ent, .cif, .mmcif, .pdbx | Plan limit, up to 50 files | Structures fitted to the reference |
PDB IDs can be fetched directly from RCSB. Fetched entries use PDBx/mmCIF, which preserves long chain identifiers and structures that cannot be represented in legacy PDB format.
In Structure mode, atoms correspond only when all of these identifiers match:
chain ID + author residue number + insertion code + record identity + atom nameFor protein ATOM records, the residue name is not part of the key, so atoms shared by a point mutation can still be compared. For HETATM records, record identity includes the residue name. A ligand atom in LIG therefore cannot collide with a protein atom or an atom from a differently named heterogen at the same author position.
In Chain mode, the calculator first separates the structures into chains and then compares every reference chain with every comparison chain. Chain ID is omitted from the atom key inside each selected pair, which allows chain A to be compared with an otherwise corresponding chain B.
Alternate coordinate locations are not counted twice. The calculator keeps blank shared atoms, chooses one occupancy-preferred alternate conformation per residue, and reports removed duplicate records as a warning. Atom pairs are never discarded as outliers after fitting, so every reported matched atom contributes to the RMSD.
This is an identifier-based RMSD calculator, not a sequence or structural alignment search. If chain IDs, numbering, or sequences differ substantially, use USAlign, which finds a structural correspondence before reporting RMSD and TM-score.
| Setting | Options | Default | What it controls |
|---|---|---|---|
Atom selection | Alpha carbons, Backbone, All protein atoms, All heavy atoms, All coordinate records | Alpha carbons | Atoms included in the reported RMSD |
Alignment atom selection | Same choices, or Same as RMSD selection | Same as RMSD selection | Atoms used to calculate the fitted rotation and translation |
Compare by | Structure, Chain | Structure | One structure-level result or every reference-chain versus comparison-chain pair |
RMSD weighting | Uniform, Atomic mass | Uniform | Equal atom contribution or contribution proportional to atomic mass |
Multi-model structures | First model only, All model pairs | First model only | Whether multi-model PDB or mmCIF files contribute one model or every model pair |
Residue numbers | Author positions and inclusive ranges, such as 10:50,75,100:120 | All residues | Limits both structures to selected author residue numbers |
Automatic alignment | On, off | On | Fits the comparison coordinates before measuring RMSD |
Return fitted structures | On, off | On | Returns each comparison model with the reported transformation applied |
| Selection | Included records | Best use |
|---|---|---|
| Alpha carbons | CA atoms from protein ATOM records | Overall fold and backbone comparison |
| Backbone | N, CA, C, and O from protein ATOM records | More detailed main-chain comparison |
| All protein atoms | Every protein ATOM record, including protein hydrogens when present | Side-chain and full-protein comparison with matching topology |
| All heavy atoms | Non-hydrogen ATOM and HETATM records | Protein, ligand, cofactor, ion, and modified-residue comparison without hydrogen sensitivity |
| All coordinate records | Every ATOM and HETATM record | Identically prepared structures, including hydrogens and solvent |
All-heavy and all-coordinate modes require matching ligand, solvent, and modified-residue identifiers. A low matched coverage means the reported RMSD describes only the shared subset.
Atomic-mass weighting reads element symbols from the coordinate records. If an element is missing or unsupported, the comparison stops with an error rather than assigning an arbitrary mass. Uniform weighting avoids that dependency and is the conventional choice for Cα and backbone RMSD.
The alignment selection can differ from the RMSD selection. For example, align on alpha carbons and measure all heavy atoms to ask how well the full atomic model agrees after the protein fold is fitted. In that case, the reported RMSD is minimized for the Cα fit selection, not necessarily for the heavier measurement selection.
Residue numbers can restrict both fitting and measurement to a domain, binding site, or stable core. It uses author residue numbers from the coordinate files. Both structures must use the same numbers for this filter; use USAlign when their numbering differs.
| Column | Meaning |
|---|---|
Reference | Reference structure, model, and chain label where applicable |
Comparison | Comparison structure, model, and chain label |
RMSD (Å) | Weighted root mean square displacement after the selected fit |
Measured atoms | Number of matched atoms used for RMSD |
Fit atoms | Number of matched atoms used to determine the transformation |
Reference coverage (%) | Percentage of selected reference atoms that were matched |
Comparison coverage (%) | Percentage of selected comparison atoms that were matched |
Unmatched reference atoms | Selected atoms found only in the reference |
Unmatched comparison atoms | Selected atoms found only in the comparison |
Automatic alignment requires at least three non-collinear matched fit atoms and three matched measurement atoms. A comparison that cannot meet that minimum is reported as a warning instead of producing a misleading or geometrically ambiguous value.
RMSD has meaning only for the atom pairs that were actually measured. Coverage shows how representative that shared set is.
| Result pattern | Interpretation |
|---|---|
| Low RMSD, high coverage | Most selected atoms occupy similar positions after fitting |
| Low RMSD, low coverage | A small shared region agrees; the result does not establish whole-structure similarity |
| High global RMSD, localized per-residue peaks | A loop, terminus, ligand, or moving domain may dominate the global value |
| High RMSD across many residues | The selected coordinates differ broadly, or the chosen correspondence and fit region are unsuitable |
| Unequal reference and comparison coverage | One structure contains selected atoms, residues, chains, or heterogens absent from the other |
Comparisons are most defensible when they report the atom selection, fit selection, weighting, matched atom count, and both coverage values alongside RMSD. Values from different selections or different matched subsets are not directly interchangeable.
The per-residue table applies the same global transformation and then groups squared atom displacements by reference residue. This highlights flexible termini, loops, binding-site changes, and local side-chain differences that may be hidden by one global value.
Per-residue RMSD is not a separate fit for each residue. Every residue is evaluated in the coordinate frame established by the global or chain-level alignment.
| File | Contents |
|---|---|
Fitted .pdb or .cif structures | Full comparison model in Structure mode, or the selected comparison chain in Chain mode, after applying the reported rigid-body transform |
rmsd_transformations.json | Rotation matrix, translation in Å, fit selection, measurement selection, and weighting for every successful comparison |
| Spreadsheet exports | Global or per-residue result rows as CSV or Excel |
Fitted mmCIF input remains mmCIF. A PDB input with a chain ID that cannot fit the one-character legacy PDB field is also returned as mmCIF.
Each run returns at most 100 fitted coordinate files and 100 MB of fitted coordinate content. If that download limit is reached, the complete numerical, per-residue, and transformation results are still returned. Requests are also limited to 1,000 structure, model, or chain comparisons and 250,000 per-residue rows so unusually large model or chain combinations fail with an actionable message instead of exhausting the browser.
Protein structure RMSD is a coordinate-distance metric for a defined set of corresponding atoms. After superposition, it summarizes the squared distances between those atom pairs as one value in ångströms. Lower values mean closer agreement for that exact correspondence, selection, and fit, but RMSD does not discover which residues should correspond.
For corresponding atoms with weights , ProteinIQ calculates:
is the optimal proper rotation calculated from the matched fit atoms, so mirror images cannot be fitted by reflection. translates the rotated comparison into the reference coordinate frame. With uniform weighting, every is 1. With atomic-mass weighting, is the mass of the matched element.
The fit minimizes the weighted squared displacement of the selected fit atoms. A separate measurement selection is evaluated after that same transformation, which is why fitting on Cα atoms and measuring heavy atoms can produce a heavy-atom RMSD that is not itself minimal.
There is no universal RMSD threshold for structural similarity. Global RMSD is sensitive to protein size and to a few large deviations, including flexible termini, loops, and domain motion. Matched coverage, per-residue RMSD, and the biological question determine whether the global value is informative.
| Goal | Tool |
|---|---|
| One reference versus several structures with matching identifiers | RMSD calculator |
| Different residue numbering, sequences, or chain layouts | USAlign |
| Complete pairwise matrix for a topology-consistent ensemble | pyRMSD |
| Protein complex interface accuracy against a native structure | DockQ |
| RMSD across every frame of a trajectory | MD trajectory analysis |
Use Cα RMSD for overall fold comparison, backbone RMSD for main-chain geometry, and heavy or all-coordinate RMSD only when the structures have closely matching atomic composition. Compare values only when atom selection, fitting, weighting, and coverage are consistent.
The structures differ in chain IDs, author residue numbering, insertion codes, missing atoms, or atom names. Chain mode can reveal corresponding chains with different IDs. For broader correspondence problems, use USAlign.
The warning panel states the reason. Common causes are an invalid coordinate file, no matching selected atoms, fewer than three matching atoms, or collinear fit atoms that cannot define a unique three-dimensional rotation.
Yes, when the ligand is present in both structures with matching chain, residue, insertion-code, and atom-name identifiers. Choose All heavy atoms and use chain mode when the ligand has its own chain. Symmetry-corrected ligand RMSD is not applied, so chemically equivalent atom permutations require a docking-specific evaluation workflow.
For alternate locations, one occupancy-preferred residue conformer is selected and duplicate coordinates are removed. Multi-model files use model 1 by default; All model pairs evaluates every reference model against every comparison model.
Disable it only when coordinates are already in the same reference frame and the absolute displacement is the quantity of interest. Without alignment, rigid translation or rotation contributes directly to RMSD.
Calculate pairwise RMSD matrices for PDB structure ensembles with pyRMSD, including the condensed matrix and source statistics files.
Assess docking model quality by comparing predicted complexes against native references. DockQ v2.1.3 supports protein, nucleic-acid, and supported small-molecule interfaces with faithful native metrics.
PoseBusters validates generated or docked molecular poses with chemically and structurally grounded quality checks for molecular geometry, intermolecular interactions, and optional reference-pose agreement.
Assign protein secondary structure using the DSSP algorithm. The gold standard for hydrogen bond-based structure assignment from coordinates.
Validate protein structure quality with all-atom contact analysis, Ramachandran plots, rotamer assessment, and geometry checks.
Generate a downloadable PDBsum structural summary report archive for a single protein structure.
Calculate the radius of gyration (Rg) for protein structures from PDB files. Supports multiple chains and atom selection options.
Calculate Solvent Accessible Surface Area (SASA) for protein structures using the Shrake-Rupley algorithm.
Predict metal and water binding sites in protein structures using 3D convolutional neural networks (AllMetal3D + Water3D).
Scoring function for interprotein interactions in AlphaFold2, AlphaFold3 and Boltz predictions. Calculates ipSAE, ipTM, pDockQ, pDockQ2, and LIS scores to assess protein-protein interface quality.
Upload or fetch a reference structure and one or more comparison structures, choose the atoms to measure, then select Calculate. ProteinIQ matches corresponding coordinates, performs an optimal rigid-body fit, and returns global RMSD, matched coverage, per-residue RMSD, fitted structures, and the rotation and translation applied to each comparison.
For a verified example, fetch 1UBQ as the reference and 1D3Z as the comparison. Keep the default settings:
RMSD atom selection: Alpha carbons (CA)
Alignment atom selection: Same as RMSD selection
Compare by: Structure
RMSD weighting: Uniform
Multi-model structures: First model only
Automatic alignment: OnThe current RCSB coordinate files produce:
Reference: 1UBQ
Comparison: 1D3Z:model_1
RMSD: 0.521 Å
Measured atoms: 76
Fit atoms: 76
Reference coverage: 100%
Comparison coverage: 100%1D3Z contains 10 NMR models, so first-model mode also reports that only model 1 was used. Choose All model pairs to compare every 1D3Z model.
The 0.521 Å result is reproducible for the current RCSB PDBx/mmCIF files and the settings shown above. It compares all 76 matched Cα atoms in 1UBQ with model 1 of 1D3Z; it is not a general threshold for deciding whether two unrelated proteins have the same fold.
| Input | Accepted formats | Limit | Matching role |
|---|---|---|---|
| Reference structure | .pdb, .ent, .cif, .mmcif, .pdbx | One file, up to 50 MB | Fixed coordinate set |
| Comparison structures | .pdb, .ent, .cif, .mmcif, .pdbx | Plan limit, up to 50 files | Structures fitted to the reference |
PDB IDs can be fetched directly from RCSB. Fetched entries use PDBx/mmCIF, which preserves long chain identifiers and structures that cannot be represented in legacy PDB format.
In Structure mode, atoms correspond only when all of these identifiers match:
chain ID + author residue number + insertion code + record identity + atom nameFor protein ATOM records, the residue name is not part of the key, so atoms shared by a point mutation can still be compared. For HETATM records, record identity includes the residue name. A ligand atom in LIG therefore cannot collide with a protein atom or an atom from a differently named heterogen at the same author position.
In Chain mode, the calculator first separates the structures into chains and then compares every reference chain with every comparison chain. Chain ID is omitted from the atom key inside each selected pair, which allows chain A to be compared with an otherwise corresponding chain B.
Alternate coordinate locations are not counted twice. The calculator keeps blank shared atoms, chooses one occupancy-preferred alternate conformation per residue, and reports removed duplicate records as a warning. Atom pairs are never discarded as outliers after fitting, so every reported matched atom contributes to the RMSD.
This is an identifier-based RMSD calculator, not a sequence or structural alignment search. If chain IDs, numbering, or sequences differ substantially, use USAlign, which finds a structural correspondence before reporting RMSD and TM-score.
| Setting | Options | Default | What it controls |
|---|---|---|---|
Atom selection | Alpha carbons, Backbone, All protein atoms, All heavy atoms, All coordinate records | Alpha carbons | Atoms included in the reported RMSD |
Alignment atom selection | Same choices, or Same as RMSD selection | Same as RMSD selection | Atoms used to calculate the fitted rotation and translation |
Compare by | Structure, Chain | Structure | One structure-level result or every reference-chain versus comparison-chain pair |
RMSD weighting | Uniform, Atomic mass | Uniform | Equal atom contribution or contribution proportional to atomic mass |
Multi-model structures | First model only, All model pairs | First model only | Whether multi-model PDB or mmCIF files contribute one model or every model pair |
Residue numbers | Author positions and inclusive ranges, such as 10:50,75,100:120 | All residues | Limits both structures to selected author residue numbers |
Automatic alignment | On, off | On | Fits the comparison coordinates before measuring RMSD |
Return fitted structures | On, off | On | Returns each comparison model with the reported transformation applied |
| Selection | Included records | Best use |
|---|---|---|
| Alpha carbons | CA atoms from protein ATOM records | Overall fold and backbone comparison |
| Backbone | N, CA, C, and O from protein ATOM records | More detailed main-chain comparison |
| All protein atoms | Every protein ATOM record, including protein hydrogens when present | Side-chain and full-protein comparison with matching topology |
| All heavy atoms | Non-hydrogen ATOM and HETATM records | Protein, ligand, cofactor, ion, and modified-residue comparison without hydrogen sensitivity |
| All coordinate records | Every ATOM and HETATM record | Identically prepared structures, including hydrogens and solvent |
All-heavy and all-coordinate modes require matching ligand, solvent, and modified-residue identifiers. A low matched coverage means the reported RMSD describes only the shared subset.
Atomic-mass weighting reads element symbols from the coordinate records. If an element is missing or unsupported, the comparison stops with an error rather than assigning an arbitrary mass. Uniform weighting avoids that dependency and is the conventional choice for Cα and backbone RMSD.
The alignment selection can differ from the RMSD selection. For example, align on alpha carbons and measure all heavy atoms to ask how well the full atomic model agrees after the protein fold is fitted. In that case, the reported RMSD is minimized for the Cα fit selection, not necessarily for the heavier measurement selection.
Residue numbers can restrict both fitting and measurement to a domain, binding site, or stable core. It uses author residue numbers from the coordinate files. Both structures must use the same numbers for this filter; use USAlign when their numbering differs.
| Column | Meaning |
|---|---|
Reference | Reference structure, model, and chain label where applicable |
Comparison | Comparison structure, model, and chain label |
RMSD (Å) | Weighted root mean square displacement after the selected fit |
Measured atoms | Number of matched atoms used for RMSD |
Fit atoms | Number of matched atoms used to determine the transformation |
Reference coverage (%) | Percentage of selected reference atoms that were matched |
Comparison coverage (%) | Percentage of selected comparison atoms that were matched |
Unmatched reference atoms | Selected atoms found only in the reference |
Unmatched comparison atoms | Selected atoms found only in the comparison |
Automatic alignment requires at least three non-collinear matched fit atoms and three matched measurement atoms. A comparison that cannot meet that minimum is reported as a warning instead of producing a misleading or geometrically ambiguous value.
RMSD has meaning only for the atom pairs that were actually measured. Coverage shows how representative that shared set is.
| Result pattern | Interpretation |
|---|---|
| Low RMSD, high coverage | Most selected atoms occupy similar positions after fitting |
| Low RMSD, low coverage | A small shared region agrees; the result does not establish whole-structure similarity |
| High global RMSD, localized per-residue peaks | A loop, terminus, ligand, or moving domain may dominate the global value |
| High RMSD across many residues | The selected coordinates differ broadly, or the chosen correspondence and fit region are unsuitable |
| Unequal reference and comparison coverage | One structure contains selected atoms, residues, chains, or heterogens absent from the other |
Comparisons are most defensible when they report the atom selection, fit selection, weighting, matched atom count, and both coverage values alongside RMSD. Values from different selections or different matched subsets are not directly interchangeable.
The per-residue table applies the same global transformation and then groups squared atom displacements by reference residue. This highlights flexible termini, loops, binding-site changes, and local side-chain differences that may be hidden by one global value.
Per-residue RMSD is not a separate fit for each residue. Every residue is evaluated in the coordinate frame established by the global or chain-level alignment.
| File | Contents |
|---|---|
Fitted .pdb or .cif structures | Full comparison model in Structure mode, or the selected comparison chain in Chain mode, after applying the reported rigid-body transform |
rmsd_transformations.json | Rotation matrix, translation in Å, fit selection, measurement selection, and weighting for every successful comparison |
| Spreadsheet exports | Global or per-residue result rows as CSV or Excel |
Fitted mmCIF input remains mmCIF. A PDB input with a chain ID that cannot fit the one-character legacy PDB field is also returned as mmCIF.
Each run returns at most 100 fitted coordinate files and 100 MB of fitted coordinate content. If that download limit is reached, the complete numerical, per-residue, and transformation results are still returned. Requests are also limited to 1,000 structure, model, or chain comparisons and 250,000 per-residue rows so unusually large model or chain combinations fail with an actionable message instead of exhausting the browser.
Protein structure RMSD is a coordinate-distance metric for a defined set of corresponding atoms. After superposition, it summarizes the squared distances between those atom pairs as one value in ångströms. Lower values mean closer agreement for that exact correspondence, selection, and fit, but RMSD does not discover which residues should correspond.
For corresponding atoms with weights , ProteinIQ calculates:
is the optimal proper rotation calculated from the matched fit atoms, so mirror images cannot be fitted by reflection. translates the rotated comparison into the reference coordinate frame. With uniform weighting, every is 1. With atomic-mass weighting, is the mass of the matched element.
The fit minimizes the weighted squared displacement of the selected fit atoms. A separate measurement selection is evaluated after that same transformation, which is why fitting on Cα atoms and measuring heavy atoms can produce a heavy-atom RMSD that is not itself minimal.
There is no universal RMSD threshold for structural similarity. Global RMSD is sensitive to protein size and to a few large deviations, including flexible termini, loops, and domain motion. Matched coverage, per-residue RMSD, and the biological question determine whether the global value is informative.
| Goal | Tool |
|---|---|
| One reference versus several structures with matching identifiers | RMSD calculator |
| Different residue numbering, sequences, or chain layouts | USAlign |
| Complete pairwise matrix for a topology-consistent ensemble | pyRMSD |
| Protein complex interface accuracy against a native structure | DockQ |
| RMSD across every frame of a trajectory | MD trajectory analysis |
Use Cα RMSD for overall fold comparison, backbone RMSD for main-chain geometry, and heavy or all-coordinate RMSD only when the structures have closely matching atomic composition. Compare values only when atom selection, fitting, weighting, and coverage are consistent.
The structures differ in chain IDs, author residue numbering, insertion codes, missing atoms, or atom names. Chain mode can reveal corresponding chains with different IDs. For broader correspondence problems, use USAlign.
The warning panel states the reason. Common causes are an invalid coordinate file, no matching selected atoms, fewer than three matching atoms, or collinear fit atoms that cannot define a unique three-dimensional rotation.
Yes, when the ligand is present in both structures with matching chain, residue, insertion-code, and atom-name identifiers. Choose All heavy atoms and use chain mode when the ligand has its own chain. Symmetry-corrected ligand RMSD is not applied, so chemically equivalent atom permutations require a docking-specific evaluation workflow.
For alternate locations, one occupancy-preferred residue conformer is selected and duplicate coordinates are removed. Multi-model files use model 1 by default; All model pairs evaluates every reference model against every comparison model.
Disable it only when coordinates are already in the same reference frame and the absolute displacement is the quantity of interest. Without alignment, rigid translation or rotation contributes directly to RMSD.
Calculate pairwise RMSD matrices for PDB structure ensembles with pyRMSD, including the condensed matrix and source statistics files.
Assess docking model quality by comparing predicted complexes against native references. DockQ v2.1.3 supports protein, nucleic-acid, and supported small-molecule interfaces with faithful native metrics.
PoseBusters validates generated or docked molecular poses with chemically and structurally grounded quality checks for molecular geometry, intermolecular interactions, and optional reference-pose agreement.
Assign protein secondary structure using the DSSP algorithm. The gold standard for hydrogen bond-based structure assignment from coordinates.
Validate protein structure quality with all-atom contact analysis, Ramachandran plots, rotamer assessment, and geometry checks.
Generate a downloadable PDBsum structural summary report archive for a single protein structure.
Calculate the radius of gyration (Rg) for protein structures from PDB files. Supports multiple chains and atom selection options.
Calculate Solvent Accessible Surface Area (SASA) for protein structures using the Shrake-Rupley algorithm.
Predict metal and water binding sites in protein structures using 3D convolutional neural networks (AllMetal3D + Water3D).
Scoring function for interprotein interactions in AlphaFold2, AlphaFold3 and Boltz predictions. Calculates ipSAE, ipTM, pDockQ, pDockQ2, and LIS scores to assess protein-protein interface quality.