rhofit test on Wlodek et al. (2006) test set.


  1. introduction and methods
  2. rhofit results
  3. comparison between rhofit and FitMAP results
  4. autobuster refinements from rhofit best fit ligands
  5. test of rhofit -scanchirals: can rhofit work from ligand with scrambled chiral centers

1. introduction and methods

rhofit is designed to flexibly fit ligand molecules into difference density. The AFITT program by OpenEye Scientific Software has similar capabilities. In the paper describing the method Wlodek et al. (2006) Acta Cryst. D 62:741-749, 11 protein-ligand complexes are used as a test of the procedure (see Tables 1 and 2). It is found that the procedure can refit ligands into the 2Fo-Fc density map from the model refined with the ligand with a maximum rmsd to the pdb position of 0.8Å.

To test that BUSTER with rhofit can cope with these ligands it was decided to apply the procedures on the Wlodek et al. (2006) test set. Rather than directly using the 2Fo-Fc density map from the refined model the following procedure was adopted:

2. rhofit results

PDB code ligand code rhofit ligand rms disp to pdb position picture (1)
1di9 MSQ 0.55 1di9.png
1a28 STR A site 0.12 1a28_a.png
1a28 STR B site 0.15 1a28_b.png
1xqd DND 0.36 1xqd.png
1cbs REA 0.56 1cbs.png
1ld8 U49 0.31 1ld8.png
1obd ATP 0.41 1obd.png
1pzp FTA 300 site 1.39 (2) 1pzp_300.png
1pzp FTA 301 site 0.48 1pzp_301.png
1ajx AH1 0.29 1ajx.png
1err RAL A site 0.39 1err_a.png
1err RAL B site 0.48 1err_b.png
1b0f SEI 0.53 (3) 1b0f.png
1ibw PVH (4) -
    1. Click on the picture to expand (or right click to get in another tab)! Difference density as found by autoBUSTER refine -L contoured at 3.5 sigma in green, rhofit best solution in atom-colored ball and stick. pdb ligand position in purple lines.
    2. In this case only autoBUSTER refine -L failed to identify this weak binding site. An autoBUSTER refine -Lpdb was then used with the binding site being manually defined by the waters inserted into it in the original run.
    3. The SEI ligand is covalently bound to the elastase protein via and ester bond to the active site serine residue. This version of rhofit does not explicitly consider covalent ligand protein bonds. Because of this the ligand was considered as unbound with the serine CB and OG atoms being removed from the fixed protein model in the rhofit run. The rhofit best solution places the ligand with atom C71 adjacent (1.0Å) to the active site serine 195 OG atom to which it is bonded. In the picture of the solution the serine side chain is shown in orange and pink. It can be noted that the geometry of the link would be improved by sensible restraints, but the same is true of 1b0f.pdb!
    4. The PVH "ligand" is a histidine residue covalently bound to the C terminus of the protein, with its carboxylic acid esterified. There are 3 NCS copies and at 3.2Å resolution the density for this is a blob. It is simply not sensible to model this residue without taking into account the its bonds.

3. comparison between rhofit and FitMAP results

Comparison between rhofit to FitMAP is shown in the charts:

4. autobuster refinements from rhofit best fit ligands

to be written up

5. Test of rhofit -scanchirals: can rhofit work from ligand with scrambled chiral centers?

PDB ligand code # chirals rhofit -scanchirals identifies same chiral comination as pdb ligand rmsd msdchem rhofit best to scrambled rhofit best
1a28 STR 6 yes 0.13
1xqd DND 8 yes (1) 0.04
1ld8 U49 1 no (2) 0.36
1obd ATP 4 yes 0.06
1ajx AH1 4 yes 0.16
1b0f SEI 3 yes 0.32
    1. for 8 chiral centres a divide and conquer strategy was used where the correct chiralities were determined for the two sugar rings were determined separately.
    2. chiral center is pretty much flat in pdb. From the -scanchiral runs I think it likely that msdchem has the incorrect chirality from the center.