Parent directory: 8_symm
Unmerged data: hemh_unmerged.mtz
Target sequence: hemh.fasta
Homologue structure: 1ak1.pdb

1. Introduction

This example illustrates space group assignment in the presence of pseudosymmetry and twinning.

The crystal structure of Ferrochelatase-1 (HemH) from Bacillus anthracis (PDB code 2c8j) presented a moderately difficult case for space group assignment. The lattice parameters, merging statistics and systematic absences favoured the space group P21212. A molecular replacement solution with a high contrast was found using a search model generated from the PDB entry 1ak1 (sequence identity of 73%). However, it was not possible to refine this solution (the lowest value of R-free was about 40%). The structure has been eventually solved, but in a monoclinic space group. In this tutorial we follow the whole procedure including revision of space group assignment.

2. Input data

The file hemh_unmerged.mtz contains unmerged HemH data presented in the space group P222. As these are unmerged data, the space group assignment only affects the order and indexing of observations and can be altered with no loss of information using pointless (the first program in aimless pipeline).

The file 1ak1.pdb was downloaded from the PDB and represents a crystal structure of Ferrochelatase-1 from Bacillus subtilis containing one protein molecule in the asymmetric unit. This structure will be used as a search model for molecular replacement.

3. Structure solution

  1. Run aimless pipeline to identify bad images and resolution cut-off
  2. Rerun aimless pipeline with bad data removed
  3. Try to solve the structure using molecular replacement, model 1ak1, and merged data from (2)
  4. Refine the structure (3) and take note of R and R-free, examine the refined model vs. electron density maps. Is there any potential for model improvement?
  5. Run aimless pipeline selecting the second hit
  6. Solve the structure using molecular replacement, model 1ak1, and merged data from (5)
  7. Refine structure from (6), examine model vs. map, compare refinement statistics with one from step (4)