[phenixbb] Overlapping ligand-protein side chain density

Phan, Jason jason.phan at vanderbilt.edu
Wed Feb 28 18:06:10 PST 2018

Thank you, Dorothee & Henry for the very informative articles. I agree that their occupancies don’t have to add up to unity since they’re separate entities.

I’d encountered scenario 7 before with two doubly disordered side chains with one conformer each sharing the same space. The altloc and constrained group occupancies combination worked just as described. However, this time, a different protein, with a ligand and a side chain sharing the same space, group occupancy constraints do not work. The ligand and the PHE residue each has just a single conformation as far as I can tell. When the ligand is bound, the PHE side chain must be out, thus the PHE must have at least two conformations for this to be physically possible. I followed the protocol in the News Letter, created an alternative conformation for the phenylanine in question and constrained the group occupancies accordingly:

refinement {
   refine {
      occupancies {
         constrained_group {
             selection = resname LIG or (chain B and resseq 890 and altloc A)

The ligand and APHE got pushed away from one another and off of their respective densities. Occupancies for both were reduced to 0.5 for the refinement. What am I doing wrong here? Does the ligand need to have an altloc as well? But I don’t see an alternative conformation.


On Feb 28, 2018, at 5:35 PM, van den Bedem, Henry <vdbedem at slac.stanford.edu<mailto:vdbedem at slac.stanford.edu>> wrote:

If the phe is a gatekeeper, shouldn’t the ligand be refined at partial occupancy; i.e. occupancies not necessarily have to ‘add up’ as you suggest? Maybe this link is helpful too: www.biorxiv.org/content/early/2018/01/24/253419<http://www.biorxiv.org/content/early/2018/01/24/253419>


From: <phenixbb-bounces at phenix-online.org<mailto:phenixbb-bounces at phenix-online.org>> on behalf of "Phan, Jason" <jason.phan at vanderbilt.edu<mailto:jason.phan at vanderbilt.edu>>
Date: Wednesday, February 28, 2018 at 1:30 PM
To: "phenixbb at phenix-online.org<mailto:phenixbb at phenix-online.org>" <phenixbb at phenix-online.org<mailto:phenixbb at phenix-online.org>>
Subject: [phenixbb] Overlapping ligand-protein side chain density


A piece of a ligand and the side chain of a “gate-keeper” Phe occupy the same space with well-defined features observed for both (resolution is 1.7 A). It looks about 50:50. How do you refine both ligand and protein side chain in this case? A couple of phenixbb suggestions for dealing with ligand-ligand overlapping density have been considered. With the first suggestion, the two entities still clashed and moved apart off of their respective densities. The second suggestion is not applicable in this case since the other molecule is not a ligand but part of the protein but it was tested anyway. Although there was no bumping, the occupancies don’t add up, resulting in a big blob of negative density around the ligand piece.

Only two comments:

  1.  At that resolution, constrained group occupancy refinement should work reasonably well (provided you can model the 2 entities). Then you also do not have clashes between the molecules, because Occ(A)+Occ(B)=1, meaning when one (A) is there, the other one (B) is not. This works with refmac (external keyword file); if you need more sophisticated occupancy re/constraints SHELXL may offer more opportunities.
  2.  There is no necessity for the two NCS copies of the binding site to look exactly the same (non-equivalent). Maybe there is a good reason/story (accessibility, contacts etc) for one site to be occupied differently than the other one.

Best, BR
Modeling two molecules that occupy overlapping binding sites in a structure simply involves designating them as alternate conformers, with the same chain and residue number, and an occupancy that sums to 1.0. For example, if you have an AMP and an ADP that occupy the same binding site, you would define them as

AAMP B 501
BADP B 501

and initially set the occupancies for the atoms in each conformer to the ratio (50:50, 30:70, etc.) that you observe in the density.

Refinement in this manner is straightforward in PHENIX.


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