Tool: ESMFold

ESMFold (Evolutionary Scale Modeling) is an artificial intelligence method for predicting protein structures. The method is described in:

Evolutionary-scale prediction of atomic-level protein structure with a language model. Lin Z, Akin H, Rao R, Hie B, Zhu Z, Lu W, Smetanin N, Verkuil R, Kabeli O, Shmueli Y, Dos Santos Costa A, Fazel-Zarandi M, Sercu T, Candido S, Rives A. Science. 2023 Mar 17;379(6637):1123-1130.

The ChimeraX ESMFold tool:

• finds and retrieves existing models from the ESM Metagenomic Atlas, which contains over 600 million predicted protein structures
  
  
• runs new ESMFold predictions on the prediction server provided by the ESM Metagenomic Atlas

ESMFold-predicted structures vary in confidence levels (see coloring) and should be interpreted with caution. The related tool ESMFold Error Plot plots residue-residue alignment errors for ESMFold structures. The ESMFold tool is also implemented as the esmfold command. See the ChimeraX ESMFold example. See also: AlphaFold, Blast Protein, Modeller Comparative, Model Loops, Rotamers

ESMFold Dialog
ESMFold Coloring Dialog
ESM Fold Error Plot

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ESMFold Dialog

The ESMFold tool can be opened from the Structure Prediction section of the Tools menu and manipulated like other panels (more...).

The Sequence can be specified by UniProt name or accession number, pasted in as plain text, or chosen from the menu of currently open protein structure chains.

Fetch
Search
Predict
Options

← Fetch gets the most sequence-similar model available from the ESM Metagenomic Atlas for each specified chain. Specifying a whole model specifies all of its protein chains. For each chain, a model is obtained for the single top hit identified by K-mer search of the ESM Metagenomic Atlas. The corresponding command is esmfold match. If the sequence was specified by structure chain, then:

1. the chain ID of the predicted structure is made the same as the corresponding chain of the existing model
2. the predicted structure is superimposed onto the existing chain using matchmaker, and the following are reported in a table in the Log:
   • Chain – chain ID in ChimeraX
   • MGnify Id – sequence ID used by the ESM Metagenomic Atlas
   • RMSD – Cα root-mean-square deviation between the predicted and experimental structures, over all residues of the latter
   • Length – number of residues in the predicted structure
   • Seen – number of residues with atomic coordinates in the experimental structure
   • % Id – percent identity in the sequence alignment generated by matchmaker for superposition; the number of positions with identical residues divided by the length of the shorter sequence
3. by default, the predicted structure is trimmed to the same residue range as the existing chain (details...)
4. the following attributes are assigned to the residues of the predicted structure:
   • c_alpha_distance – Cα distance between corresponding positions of the predicted and existing chains after their superposition (step 2 above)
   • missing_structure – positions missing from the coordinates of the existing chain
   • same_sequence – positions with different residue types than the existing chain
   These attributes can be used for coloring and other purposes.

The fetched models are stored locally in ~/Downloads/ChimeraX/ESMFold/, where ~ indicates a user's home directory. If a file specified for opening is not found in this local cache, the file will be fetched and cached.

← Search uses a BLAST web service hosted by the UCSF RBVI to search the ESM Metagenomic Atlas using default parameters: BLOSUM62 amino acid similarity matrix for scoring the hits, similarity score cutoff e-value 1e-3, returning a maximum of 100 of unique sequences. However, different values of these parameters can be specified using the corresponding command, esmfold search. Search differs from Fetch in that it uses BLAST instead of fast (but low-sensitivity) K-mer searching, accepts only a single chain or sequence as input, and returns a list of hits for the user to inspect, rather than fetching the single top hit per chain automatically. When results are returned, the hits are listed in a Blast Protein window. Double-clicking a hit uses esmfold fetch to retrieve the model, or multiple chosen hits can be retrieved at once by using the results panel context menu or Load Structures button (details...).

← Predict runs a calculation on the prediction server provided by the from the ESM Metagenomic Atlas. The corresponding command is esmfold predict.

← The Options button shows/hides additional options:

• Results directory (default ~/Downloads/ChimeraX/ESMFold/) – the pathname (name and location) of a folder or directory in which to store prediction results. Clicking Browse brings up a file browser window for choosing it interactively. The directory does not need to exist already, as it will be created by running the prediction.
• Trim fetched structure to the aligned structure sequence (default on) – whether to trim a fetched structure to the same residue range as the structure to which it is aligned (details...)

The model for a sequence that was specified by structure chain will be superimposed on that chain and assigned structure-comparison attributes for further analysis (details...).

Please note the following caveats of running a prediction:

• ESMFold is faster but often less accurate than AlphaFold.
• The prediction server has a maximum sequence length of 400 residues. (The atlas contains longer predictions, up to 1024 residues.)
• No multimer prediction. Only single-chain structures (not multimers) are predicted, although structures of the individual protein chains in a complex can be predicted separately.
• PAE not available from server. Although ESMFold computes predicted aligned error (PAE), the prediction server does not provide it. The PAE is available for entries fetched from the atlas, however.
• The server developers request that users run only one prediction at a time due to capacity limitations.
• The server may time out during a prediction.

Coloring shows the ESMfold Coloring dialog for applying different color schemes to the predicted structures, as well as hiding, showing, and selecting their residues based on attribute value.

Error plot draws the ESMFold Error Plot, in which color gradations show (for each pairwise combination of residues) the expected error in position of one residue when the true and predicted structures are aligned based on the other residue.

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ESMFold Coloring Dialog

Clicking the Coloring button on the main ESMFold tool shows the ESMFold Coloring dialog for applying different color schemes to the predicted structures, as well as hiding, showing, and selecting their residues based on attribute value.

When first opened, ESMFold-predicted structures are automatically colored by the pLDDT confidence measure (same as for AlphaFold except mapped to 0-1 instead of 0-100) in the B-factor field:

• 1.0 to 0.9 – high accuracy expected
• 0.9 to 0.7 – backbone expected to be modeled well
• 0.7 to 0.5 – low confidence, caution
• 0.5 to 0.0 – should not be interpreted, may be disordered

...in other words, using

color bfactor palette esmfold

key red:low orange: yellow: cornflowerblue: blue:high  [other-key-options]

In the ESMFold Coloring dialog, the Residues to act on are specified by using the menus to choose an ESMFold-predicted model and one of the following:

• all – all residues
• confidence below [N] – based on the bfactor atom attribute (the confidence value is read from the B-factor field of the PDB file)
• C-alpha distance greater than [d] – based on the c_alpha_distance residue attribute of ESMFold models fetched by existing structure chain; Cα distance between corresponding positions of the predicted and existing chains after their automatic superposition
• missing structure – based on the missing_structure residue attribute of ESMFold models fetched by existing structure chain; positions missing from the coordinates of the existing chain
• different sequence – based on the same_sequence residue attribute of ESMFold models fetched by existing structure chain; positions with different residue types than the existing chain
• confidence above [N] – based on the bfactor atom attribute (the confidence value is read from the B-factor field of the PDB file)
• C-alpha distance less than [d] – based on the c_alpha_distance residue attribute of ESMFold models fetched by existing structure chain; Cα distance between corresponding positions of the predicted and existing chains after their automatic superposition
• paired structure – based on the missing_structure residue attribute of ESMFold models fetched by existing structure chain; positions present in the coordinates of the existing structure chain
• same sequence – based on the same_sequence residue attribute of ESMFold models fetched by existing structure chain; positions with identical residue types as the existing chain

Buttons act on the designated residues:

• Color buttons:
  • Custom for choosing a color interactively using the system color picker
  • a series of square buttons for specific colors:
• Hide – hide the specified residues
• Show – show the specified residues
• Select – select the specified residues

The ESMFold Coloring dialog does not color continuously along a gradient to show the attribute values. For coloring along a gradient, see Render by Attribute and/or the commands color bfactor (for the confidence value, which is read from the B-factor field of the PDB file) and color byattribute (for other numerical attributes).

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ESMFold Error Plot

Besides the per-residue pLDDT confidence measure, ESMFold gives for each pair of residues (X,Y) the expected position error at residue X if the predicted and true structures were aligned on residue Y. These residue-residue “predicted aligned error” or PAE values are not provided by the prediction server, but are available for structures already in the ESM Metagenomic Atlas and can be shown with ESMFold Error Plot, which can be opened from the Structure Prediction section of the Tools menu and manipulated like other panels (more...). See also: esmfold contacts

Choosing the corresponding ESMFold structure from the menu of open atomic models associates it with the plot. This association allows coloring the structure as described below, and for selections on the plot to highlight the corresponding parts of the structure.

The PAE values can be either:

• fetched from the ESM Metagenomic Atlas by giving the MGnify identifier of the sequence (e.g., as evident from the name of the corresponding previously fetched ESMfold structure model)
• a json file from ESMFold (e.g., obtained by URL such as https://api.esmatlas.com/fetchConfidencePrediction/MGYP002537940442)

The PAE plot can also be shown by clicking the Error plot button on the ESMFold dialog or by using the command esmfold pae, or the command esmfold fetch or esmfold match with the option pae true.

When the mouse cursor is over the plot, the residue pair and PAE value at its current position are reported in the bottom right corner of the window.

Clicking Color PAE Domains clusters the residues into coherent domains (sets of residues with relatively low PAE values) and uses randomly chosen colors to distinguish these domains in the structure (details...). Clicking Color pLDDT returns the structure to the default confidence coloring.

The plot's context menu includes:

• Dragging box colors structure (initial default checked on) – whether dragging a box on the plot highlights the corresponding parts of the 3D structure with bright colors and makes everything else gray; if this option is unchecked, highlighting will be done with selection instead of coloring
• Color plot from structure – color the plot to match the 3D structure where the pair of residues represented by an X,Y point have the same ribbon color; show the rest of the plot in shades of gray
• Color plot rainbow – use the pae palette (default) to color the plot, with colors assigned to values as follows:

  0

  5

  10

  15

  20

  25

  30

• Color plot green – use the paegreen palette to color the plot:

  0

  5

  10

  15

  20

  25

  30

• Show chain divider lines (initial default checked on) – for multimer predictions, draw lines on the plot demarcating the end of one chain and the start of another; the lines may obscure a few chain-terminal residues in the plot, and can be hidden if this is problematic
• Save image – save the plot as a PNG file

The Color Key graphical interface or a command can be used to draw (in the main graphics window) a color key for the PAE plot. For example, to make a color key that matches the pae or paegreen scheme, respectively:

key pae :0 : : :15 : : :30  showTool true
key paegreen :0 : : :15 : : :30  showTool true

A title for the color key (e.g., “Predicted Aligned Error (Å)”) would need to be created separately with 2dlabels.