Thank you guys so much for your suggestions!
Yes, I think it's true that "if ligand density is strong enough it may not be masked by the bulk-solvent completely". In my case,
I have two different ligands in the protein-ligand structure, and using the method I described in the first email, one of the ligand has good, continuous green density but for the other ligand, as I described before, contoured at 2σ, the Fo-Fc map generated show green density for most part of the ligand, but for several atoms in the middle of the ligand, the green density is missing...

Hi Nat, you mentioned that
1. "I thought phenix.refine ignores zero-occupancy atoms when calculating the bulk solvent mask?  In fact there is an option specifically to toggle this option, and I've noticed very different results with and without it."

I am wondering how this could be done.

2. "Restraining the zero-occupancy ligand (and perhaps adjacent residues) is strongly recommended, especially if it contains heavier elements.  (The newer versions of the phenix.refine GUI have a button in the Output tab to set this up.)  Without restraints, you run the risk of refining the surrounding model into the ligand density.  As always this is worse at lower resolution - at 2.8Å I'm not sure what to expect."

I am wondering how this could be done.

Thank you guys so much!

Best,
Wei







On Sat, Sep 13, 2014 at 9:13 PM, Pavel Afonine <pafonine@lbl.gov> wrote:

On 9/13/14 5:57 PM, Nathaniel Echols wrote:
On Sat, Sep 13, 2014 at 3:03 PM, Pavel Afonine <pafonine@lbl.gov> wrote:
In the first case the bulk-solvent mask will be set in the ligand region and therefore it will mask ligand density (bulk-solvent will be filled into the ligand region). Depending on the strength of ligand density it may be masked completely or deteriorated.
If you follow the second option you will always get positive density in ligand area. This density may correspond to bulk-solvent, ligand or mixture of both. That is there will be no simple way to differentiate whether this density arises from the ligand or bulk-solvent.

I thought phenix.refine ignores zero-occupancy atoms when calculating the bulk solvent mask?  In fact there is an option specifically to toggle this option, and I've noticed very different results with and without it.

 It is bad both ways:
  1) If you ignore zero occupancy atoms then you fill bulk-solvent into ligand region and therefore mask the ligand;
  2) If you do not ignore zero occupancy atoms then what you compute is bulk-solvent-omit map in region around atoms with zero occupancy. This means that the "green" density you are going to see may be ligand or may be bulk-solvent or may be the mixture of the two.

In fact, #1 above is better than #2 because if ligand density is strong enough it may not be masked by the bulk-solvent completely.

Pavel