Searching for contaminants with SIMBAD

Note

Data used throughout this example can be found in <ROOT>/examples/contaminant_example. If SIMBAD is part of your CCP4 installation, then the example files can be downloaded as part of the GitHub repository.

0. Command line options

Check out this page explaining the simbad-contaminant script command line options.

1. Running the SIMBAD contaminant script

In this example, the simbad-contaminant script simply takes the crystallographic data file in MTZ format, and runs the contaminant search on your local machine.

SIMBAD can be found under the Molecular Replacement menu in the CCP4i GUI:

../../_images/ccp4i_simbad.png

Opening SIMBAD will bring you to the following menu:

../../_images/ccp4i_simbad_run.png

To run the lattice search, all that is needed is an MTZ file:

../../_images/ccp4i_simbad_contaminant.png

SIMBAD Output

On starting SIMBAD a separate window will appear summarising the progress of the SIMBAD contaminant search and any results found. The window will contain up to three tabs, the contents of which are explained below:

Log File

This displays the text output by SIMBAD as it is running. Any problems or errors will be displayed here.

../../_images/ccp4i_contaminant_log.png

Contaminant Search Results

The Contaminant Search Results tab contains 5 different sections. Below you can find information about each:

Contaminant database AMORE Rotation Search Results

../../_images/ccp4i_contaminant_results.png

This shows the results from the AMORE Rotation Search carried out on the contaminant database. The columns of the table are:

  • PDB_code: The 4 letter code representing the protein in the protein data bank
  • ALPHA: Lattice parameter alpha
  • BETA: Lattice parameter beta
  • GAMMA: Lattice parameter gamma
  • CC_F: he correlation coefficient between the observed amplitudes for the crystal and the calculated amplitudes for the model
  • RF_F: The classic R factor between the observed amplitudes for the crystal and the calculated amplitudes for the model
  • CC_I: The correlation coefficient between the observed intensities for the crystal and the sum of calculated intensities for all symmetry equivalents of the model
  • CC_P: The Patterson correlation coefficient between the crystal and the model Pattersons evaluated within the defined sphere centered on the Patterson origin
  • Icp:
  • CC_F_Z_score: Z-score of CC_F peaks
  • CC_P_Z_score: Z-score of CC_P peaks
  • Number_of_rotation_searches_producing_peak: Number of rotations searches which produce each peak [out of 5]

The structures are scored by CC_F_Z_score score where a higher score is better.

Molecular Replacement Search Results

Molecular replacement is performed on the top 20 structures identified by the Contaminant Search. This section displays the results of that molecular replacement.

By default SIMBAD runs Molecular replacement using MOLREP. If run the following columns are added to the table:

  • molrep_score: MOLREP score for the Molecular Replacement solution
  • molrep_tfscore: MOLREP translation function score for the Molecular Replacement solution

Alternatively SIMBAD can run Molecular replacement using PHASER. If run the following columns are added to the table:

  • phaser_llg: PHASER Log-likelihood gain for the Molecular Replacement solution
  • phaser_tfz: PHASER Translation Function Z-score for the Molecular Replacement solution
  • phaser_rfz: PHASER Rotational Function Z-score for the Molecular Replacement solution

Following Molecular replacement, refinement is run using REFMAC. This add the following columns are added to the table:

  • final_r_fact: R-fact score for REFMAC refinement of the Molecular Replacement solution
  • final_r_free: R-free score for REFMAC refinement of the Molecular Replacement solution

Note

Typically a result with a final_r_fact and a final_r_free below 0.45 is indicative of a solution.

Additionally if there is anomalous signal in your data set SIMBAD will try to validate the quality of the molecular replacement solution using by plotting the peaks from an anomalous fourier map. If run the following columns are added to the table:

  • dano_peak_height: The highest anomalous peaks found
  • dano_z_score: DANO peak Z-score

Top 10 Contaminant Search Downloads

This section contains the refined placed model and mtz for the top 10 solutions in the contaminant database search (as ranked by final_r_free)

Note

This section may contain less than 10 solutions if for example a solution was found and the molecular replacement search ended early

Top 10 Contaminant Search Log Files

This section contains the molecular replacement and refinement logs for the top 10 solutions in the contaminant database search (as ranked by final_r_free)

Note

This section may contain less than 10 solutions if for example a solution was found and the molecular replacement search ended early

Summary

The summary tab contains three different sections. Below you can find information about each:

../../_images/ccp4i_contaminant_summary.png

SIMBAD Summary

This details the best model found by SIMBAD and reports the final_r_fact and final_r_free scores found

Best SIMBAD result Download

This section contains the refined placed model and mtz for best solution found by the lattice parameter search (as ranked by final_r_free)

Best SIMBAD result Log Files

This section contains the molecular replacement and refinement logs for best solution found by the lattice parameter search (as ranked by final_r_free)