[phenixbb] Overlapping ligand-protein side chain density

Dorothee Liebschner dcliebschner at lbl.gov
Thu Mar 1 16:04:07 PST 2018

Hi Jason,

"I did not force the occupancies to add up to 1 but the program did it

If you use constrained groups (as defined in the parameter file you quote),
the occupancy sum will be constrained to 1.

Best wishes,


On Thu, Mar 1, 2018 at 3:35 PM, Phan, Jason <jason.phan at vanderbilt.edu>

> Thank you, Edward and Oleg for the suggestions, and Pavel for the
> clarification. You need two selections in the constrained group and an
> altloc for the ligand that’s different from the protein side chain. I did
> not force the occupancies to add up to 1 but the program did it anyway.
> Phenix refined the ligand’s occupancy to 0.68 and the PHE side chain to
> 0.32.
> This is what I did:
> I did not make an alternate conformation for the PHE, so there’s no BPHE.
> I just added the lock to the side chain starting at carbon beta. So, I have
> APHE for the side chain and BLIG for all the ligand atoms and reduced their
> occupancies to < 1.
> This is the parameter file:
> refinement {
>    refine {
>       occupancies {
>          constrained_group {
>              selection = (chain B and resseq 890) and altloc A
>              selection = resname LIG and altloc B
>    }
>        }
>    }
> }
> The rationale is in one population, the ligand is not bound and the PHE
> occupies the pocket. In the other, the ligand is bound the the PHE is out
> somewhere that I cannot observe and model, thus I did not model that
> PHE-out conformation. The refinement went to completion with the
> occupancies refined to 0.68 and 0.32 for BLIG and APHE, respectively.
> R-values dropped by a few tenths of a percent. Please let me know if
> there’s a better way to do this refinement.
> @Pavel, there’s repulsion between the selections if I don’t put an altloc
> on the ligand as suggested by Oleg.
> Jason
> On Mar 1, 2018, at 4:59 PM, Pavel Afonine <pafonine at lbl.gov> wrote:
> Hi Jason,
> Oleg is correct, you can assign the ligand and PHE different non-blanc
> altlocs (say A and B). In this case PHE and ligand will not 'see' each
> other via repulsion term of restraints. Also, their occupancies will be
> refined independently and will be constrained to be within 0 and 1, but it
> will not make sure that they add up to 1. If you want them to add up to 1,
> then make a parameter file with a constrained group as described in one of
> examples here:
> "13 typical occupancy refinement scenarios and available options in
> phenix.refine"
> http://phenix-online.org/newsletter/
> <https://na01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fphenix-online.org%2Fnewsletter%2F&data=02%7C01%7Cjason.phan%40vanderbilt.edu%7C40b308da75d345eb3cf408d57fc82098%7Cba5a7f39e3be4ab3b45067fa80faecad%7C0%7C1%7C636555419887467714&sdata=5VYe7ZcJpoBjUZVV7VTStHt2sRTUn1W4z%2FztPNHyGpE%3D&reserved=0>
> You can do it in the GUI too.
> Let us know if you more questions or problems!
> Pavel
> On 3/2/18 02:57, Oleg Sobolev wrote:
> Hi Jason,
>> 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.
> Yes, the ligand also needs an altloc, and it should be different from PHE
> altloc. This way the refinement will know that these atoms don't see each
> other. You don't have to put two alternative conformations for the ligand,
> you can have just one with partial occupancy (<1.0) which will mean that
> the ligand is not always there.
> Best regards,
> Oleg Sobolev.
>> On Feb 28, 2018, at 5:35 PM, van den Bedem, Henry <
>> 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/e
>> arly/2018/01/24/253419
>> <https://na01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.biorxiv.org%2Fcontent%2Fearly%2F2018%2F01%2F24%2F253419&data=02%7C01%7Cjason.phan%40vanderbilt.edu%7C40b308da75d345eb3cf408d57fc82098%7Cba5a7f39e3be4ab3b45067fa80faecad%7C0%7C1%7C636555419887467714&sdata=%2BeF9t7TNZ72bkVoY8GqaBvuNd1EOp%2BaRRmhlcB8OZ1g%3D&reserved=0>
>> H
>> *From: *<phenixbb-bounces at phenix-online.org> on behalf of "Phan, Jason" <
>> jason.phan at vanderbilt.edu>
>> *Date: *Wednesday, February 28, 2018 at 1:30 PM
>> *To: *"phenixbb at phenix-online.org" <phenixbb at phenix-online.org>
>> *Subject: *[phenixbb] Overlapping ligand-protein side chain density
>> All,
>> 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.
>> Diana
>> ----------------------------
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