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Command: alphafold

AlphaFold is an artificial intelligence method for predicting protein structures that has been highly successful in recent tests. The alphafold command:

Both are freely available for academic and commercial use under CC BY 4.0. Users should cite:

Highly accurate protein structure prediction with AlphaFold. Jumper J, Evans R, Pritzel A, et al. Nature. 2021 Aug;596(7873):583-589.

...and for multimer prediction:

Protein complex prediction with AlphaFold-Multimer. Evans R, O'Neill M, Pritzel A, et al. bioRxiv 2021. doi: https://doi.org/10.1101/2021.10.04.463034.

The AlphaFold Database contains modeled structures for protein sequences in UniProt:

AlphaFold Protein Structure Database: massively expanding the structural coverage of protein-sequence space with high-accuracy models. Varadi M, Anyango S, Deshpande M, et al. Nucleic Acids Res. 2022 Jan 7;50(D1):D439-D444.

The predicted structures vary in confidence levels and should be interpreted with caution. The database contains structures for single chains, not complexes; assembling the individual structures into a complex may give unphysical results where parts of the chains intersect or interact poorly with one another.

The alphafold command is also implemented as the tools AlphaFold and AlphaFold Error Plot. Several ChimeraX presentations and videos show modeling with AlphaFold and related analyses. See also: blastprotein, modeller, swapaa

Getting Models from the AlphaFold Database

Usage: alphafold fetch  uniprot-id  alignTo  chain-spec [ trim  true | false ]]colorConfidence  true | false ] [ ignoreCache  true | false ] [ pae  true | false ]
Usage: alphafold match  sequence  [ search  true | false ] [ trim  true | false ] [ colorConfidence  true | false ] [ ignoreCache  true | false ] [ pae  true | false ]
Usage: alphafold search  sequence  [ matrix  similarity-matrix ] [ cutoff  evalue ] [ maxSeqs  M ]

Options

alignTo  chain-spec
Superimpose the predicted structure from alphafold fetch onto a single chain in an already-open structure, and make its chain ID the same as that chain's. See also the trim option.
colorConfidence  true | false
Whether to color the predicted structures by the pLDDT confidence measure in the B-factor field (default true):

...in other words, using

color bfactor palette alphafold

The Color Key graphical interface or a command can be used to draw a corresponding color key, for example:

key red:low orange: yellow: cornflowerblue: blue:high  [other-key-options]
ignoreCache  true | false
The fetched models are stored locally in ~/Downloads/ChimeraX/AlphaFold/, where ~ indicates a user's home directory. If a file specified for opening is not found in this local cache or ignoreCache is set to true, the file will be fetched and cached.
search  true | false
When fetching models with alphafold match, whether to search the database for the most similar sequence if the UniProt accession number for a chain is not provided in the experimental structure's input file, or is provided but not found in the AlphaFold Database (true, default). The search uses a BLAT web service hosted by the UCSF RBVI. The closest sequence match for which a models is available will be retrieved, as long as the sequence identity is at least 25%. With search false, only the experimental structure's input file will be used as a potential source of UniProt accession numbers. When present, these are given in DBREF records in PDB format and in struct_ref and struct_ref_seq tables in mmCIF.
minimize  true | false
This option allows skipping energy-minimization of the result from alphafold predict, for faster job completion and/or to avoid failures during minimization.
trim  true | false
Whether to trim a predicted protein structure to the same residue range as the corresponding experimental structure given with the alphafold match command or the alignTo option of alphafold fetch. With trim true (default unless pae true is used):

Using trim false indicates retaining the full-length models for the UniProt sequences, which could be longer. Since a PAE plot covers the full sequence, using pae true automatically sets trim to false.

Running an AlphaFold Prediction

The alphafold predict command runs AlphaFold version 2.2.0. For monomer prediction:

Usage: alphafold predict  sequenceminimize  true | false ]

The protein sequence to predict can be given as any of the following:

  1. a chain-spec corresponding to a single chain in an atomic structure open in ChimeraX
  2. the sequence-spec of a sequence in the Sequence Viewer, in the form:  alignment-ID:sequence-ID  (details...)
  3. a UniProt name or accession number
  4. plain text pasted directly into the command line

These methods specify the entire sequence only, although a truncated version could be pasted.

For multimer (protein complex) prediction:

Usage: alphafold predict  chain-specminimize  true | false ]
– or –
Usage: alphafold predict  sequence1,sequence2[,sequence3...,sequenceN] [ minimize  true | false ]

The sequences of two or more protein chains can be specified either collectively as a chain-spec (for atomic-structure chains already open in ChimeraX), or individually within a comma-separated list using any combination of specifier types #2-4 listed above. The comma-separated list should not contain any spaces. If the same protein chain occurs multiple times in the complex, its sequence should be repeated that number of times. For example, to predict a homodimer, the same sequence (or its specifier) would need to be given twice. Prediction may only be feasible for smaller complexes (details...).

AlphaFold calculations are run using Google Colab, which is provided under the Apache 2.0 license. A warning will appear saying that this Colab notebook is from github (was not authored by Google), with a button to click to run anyway. Users will need to have a Google account and to sign into it via a browser. Once that is done, the sign-in may be remembered depending on the user's browser settings; it is not kept in the ChimeraX preferences.

A single calculation may take hours to complete (see the caveats below). The free version of Colab limits jobs to 12 hours and may terminate them at shorter times at Google's discretion (see the FAQ). Those who want to run longer and/or more frequent calculations may need to sign up for one of the paid Colab plans.

The model will be opened automatically and colored by confidence value. 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...).

Caveats

AlphaFold Predicted Aligned Error (PAE)

Besides the per-residue pLDDT confidence measure, AlphaFold 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 can be shown as a 2D plot using the command alphafold pae (details below), the AlphaFold Error Plot tool, or alphafold fetch or alphafold match with the option pae true. See also the AlphaFold Error Estimates example and video.

Usage: alphafold paeuniprotId  uniprot | file  filename ) [ model-spec ] [ palette  palette ] [ range  low,high | full ] [ plot  true | false ] [ colorDomains  true | false ] [ minSize  M ] [ connectMaxPae  N ] [ cluster  resolution ] [ dividerLines  true | false ]

With alphafold pae, the matrix of PAE values can be either:

The corresponding AlphaFold structure (already open) can be given as a model-spec to associate it with the plot. This association allows coloring by domain as described below, and for selections on the plot to highlight the corresponding parts of the structure.

By default, the PAE plot is drawn when domain coloring is not done (plot is default true when colorDomains is false) and vice versa.

Setting colorDomains to true clusters the residues into coherent domains (sets of residues with relatively low pairwise PAE values) and uses randomly chosen colors to distinguish these domains in the structure. The residues are assigned an integer domain identifier (starting with 1) as an attribute named pae_domain that can be used to specify them in commands (for example, to recolor or select specific domains). Residues not grouped into any domain are assigned a pae_domain value of None. The clustering uses the NetworkX greedy_modularity_communities algorithm with parameters:

The dividerLines option (default true) indicates whether, for multimer predictions, to 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 dividerLines false can be used if this is problematic.

The plot window has buttons for coloring the associated structure. Color PAE Domains applies coloring by PAE cluster as described above. Color pLDDT returns the structure to the default confidence coloring.

The default palette for coloring the PAE plot is pae, with colors assigned to values as follows:

0 5 10 15 20 25 30

Another palette with value range suitable for PAE plots is paegreen:

0 5 10 15 20 25 30

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., “AlphaFold Predicted Aligned Error (Å)”) would need to be created separately with 2dlabels.

Pseudobonds Colored by PAE

Residue-residue PAE values can also be shown with colored pseudobonds in the predicted structure:

Usage: alphafold contacts  res-spec1  [ toResidues  res-spec2  [ flip  true | false ] [ distance  d ] [ palette  palette ] [ range  low,high | full ] [ radius  r ] [ dashes  N ] [ outputFile  pae-file ]

See the AlphaFold Contacts example and video. See also: size, style, contacts, crosslinks

A PAE plot containing the specified residues must already be shown. The PAE matrix is not symmetrical. The first specification res-spec1 gives the aligned residues, whereas toResidues res-spec2 gives the residues whose error values are reported, except that using flip true swaps the meaning of res-spec1 and res-spec2. If one set of residues is higher-confidence (lower in pLDDT) than the other, it is usually best to specify them as the aligned residues so that the coloring will show the error values of the lower-confidence set.

Omitting the toResidues option defines res-spec2 as all residues covered by the PAE plot except for those in res-spec1; however, if toResidues is omitted and res-spec1 includes all residues in the plot, res-spec2 will also be defined as all residues in the plot.

The distance option allows limiting the number of pseudobonds by only drawing them between pairs of residues with any interresidue distance ≤ d Å (default 3.0). These pseudobonds are drawn between α-carbons regardless of which atoms were within the distance cutoff.

The default palette for coloring the pseudobonds by PAE value is paecontacts, with colors assigned to values as follows:

0 5 10 15 20

Although this palette includes value-color pairs, it may be helpful to give a value range if a colors-only palette is used instead. A range can also be used to override the values in a value-color palette, instead spacing the colors evenly across the specified range.

The pseudobond stick radius (default 0.2 Å) and number of dashes (default 1, meaning a solid stick) can also be specified.

The outputFile option allows saving a list of the residue pairs (those meeting the distance criterion) and their PAE values to a plain text file. The pae-file argument is the output file pathname, enclosed in quotation marks if it includes spaces, or the word browse to specify it interactively in a file browser window.

Examples:

alphafold contacts #1
alphafold contacts /A to /B distance 8
alphafold contacts sel palette blue:red range 1,5


UCSF Resource for Biocomputing, Visualization, and Informatics / June 2022