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USAGE.md

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Using WHISCY

After it has been installed according to the instructions in the INSTALLATION.md file, you can use WHISCY to predict the binding site of a protein.

Example

Below we will give an example of how to use WHISCY to predict the binding site of the protein 1PPE.

Get an input PDB

Download the PDB file of the protein 1PPE from the RCSB PDB website.

wget https://files.rcsb.org/download/1PPE.pdb

Setup the prediction

$ whiscy_setup 1PPE.pdb E

  2024-04-22 11:43:39,469 cli_setup:201 INFO - PDB structure with chain E saved to 1PPE_E.pdb
  2024-04-22 11:43:39,494 cli_setup:208 INFO - Atom accessibility calculated to 1PPE_E.rsa
  2024-04-22 11:43:39,495 cli_setup:212 INFO - Surface and buried residues calculated
  2024-04-22 11:43:39,513 cli_setup:255 INFO - HSSP file not found, fallback to generating MSA with blastp
  2024-04-22 11:43:39,513 cli_setup:262 INFO - Running blastp via NCBI
  2024-04-22 11:44:43,926 cli_setup:264 INFO - Result stored in 1PPE_E_blast.xml
  2024-04-22 11:44:43,933 cli_setup:277 INFO - Generating MSA using MUSCLE...
  2024-04-22 11:44:44,027 cli_setup:280 INFO - Done.
  2024-04-22 11:44:44,027 cli_setup:283 INFO - Converting MSA file to Phylseq format...
  2024-04-22 11:44:44,027 cli_setup:292 INFO - 1PPE_E.phylseq file written
  2024-04-22 11:44:44,046 cli_setup:319 INFO - Protdist calculated
  2024-04-22 11:44:44,054 cli_setup:326 INFO - Conversion table file generated
  2024-04-22 11:44:44,054 cli_setup:328 INFO - Whiscy setup finished

This script will generate a set of files needed for the prediction step with whiscy. Here it is a list of the generated files in our 1PPE complex.

Follow the links to see a set of pre-generated files for the 1PPE protein.

File name Explanation
1ppe.hssp Multiple sequence alignment download from the HSSP database
1ppe.hssp.bz2 HSSP MSA file compressed
1ppe.pdb PDB file download from the Protein Data Bank
1ppe_E.pdb 1ppe.pdb parsed to select only the given chain_id
1ppe_E.rsa SASA output of freesasa in NACCESS format of 1ppe_E.pdb file
1ppe_E.fasta Sequence of 1ppe_E.pdb. Alternative residues have been removed
1ppe_E.phylseq MSA file translated from HSSP to PHYLIP format
1ppe_E.conv PDB residue numeration to FASTA sequence numeration
1ppe_E.out Output of the protdist software on 1ppe_E.pdb
1ppe_E.sur >15 % surface residue list according to sa_pred_cutoff cutoff
1ppe_E.suract >40 % surface residue list according to sa_act_cutoff cutoff
1ppe_E.lac 0-15 % accessible residue list

Make the interface prediction

WHISCY needs four input files (generated before):

  • surface_list (.sur): List of residues in the interface.
  • conversion_table (.conv),represents the mapping of the PDB file residue numeration into the FASTA sequence numeration
  • alignment_file (.phylseq) is the MSA file in PHYLIP format
  • distance_file (.out) is the output of Protdist software
$ whiscy 1PPE_E.sur 1PPE_E.conv 1PPE_E.phylseq 1PPE_E.out -o 1PPE_E.cons
  whiscy [INFO] Parsing surface list...
  whiscy [INFO] Loading conversion table...
  whiscy [INFO] Converting...
  whiscy [INFO] Initializing score calculation...
  whiscy [INFO] Calculating scores...
  whiscy [INFO] Subtracting average value ...
  whiscy [INFO] Sorting scores...
  whiscy [INFO] Writing scores...
  whiscy [INFO] Prediction written to 1PPE_E.cons

  "I’m on a whisky diet. I’ve lost three days already."
    -  Tommy Cooper

The file 1PPE_E.cons will contain the initial predictions, but we can improve them by adding the interface propensities and smoothing the surface.

$ head 1PPE_E.cons
  0.10595   N25
  0.10595   N72
  0.10595   N74
  0.10595   N79
  0.10595  N100
  0.10595  N179
  0.10572   N95
  0.10572  N115
  0.10446   S37
  0.10446   S49

Add Interface propensities

$ whiscy_consadjust 1PPE_E.cons -o 1PPE_E.acons

consadjust [INFO] Reading input files
consadjust [INFO] Subtracting average value...
consadjust [INFO] Sorting scores...
consadjust [INFO] Writing scores...

Smooth the surface

$ whiscy_resdist 1PPE_E.pdb 1PPE_E.conv 1PPE_E.rd

residue_distance [INFO] Reading conversion table
residue_distance [INFO] Reading PDB structure from 1PPE_E.pdb
residue_distance [INFO] Residue distances written to 1PPE_E.rd

$ whiscy_parasmooth 1PPE_E.acons 1PPE_E.cons 1PPE_E.rd -o 1PPE_E.pscons

parasmooth [INFO] Reading input files
parasmooth [INFO] Calculating parameter smoothing
parasmooth [INFO] Result written to 1PPE_E.pscons

1PPE_E.pscons will contain the final predictions! 🎉

head 1PPE_E.pscons
 0.16475   V75
 0.14702   I73
 0.14633   L114
 0.14547   F82
 0.14297   V76
 0.12907   R117
 0.12127   Q81
 0.11694   N115
 0.11630   N72
 0.11512   N25

Visualizing the results

You can visualize the predictions in the input structure using whiscy_bfactor, this script will add the WHISCY scores to the B-factor column of the PDB file.

$ whiscy_bfactor 1PPE_E.pdb 1PPE_E_whiscy.pdb 1PPE_E.pscons

1PPE_E_whiscy.pdb PDB file with WHISCY scores in B-factor column has been created

Now open the structure in your favorite molecular viewer and check the WHISCY scores in the B-factor column.