Jacob Once again Boaz is leading you in the correct direction. You did send me a *.metals.edits file. You just need to change the ideal distances in this file. On another point, I ran a refinement with your inputs and the values don't seem a long way off. However, I'd wait for Pavel to take a look because the density around the Calcium ion itself seems unusual. Cheers Nigel --- Nigel W. Moriarty Building 33R0349, Physical Biosciences Division Lawrence Berkeley National Laboratory Berkeley, CA 94720-8235 Phone : 510-486-5709 Email : [email protected] Fax : 510-486-5909 Web : CCI.LBL.gov On Wed, Dec 30, 2015 at 6:27 AM, Boaz Shaanan wrote:

Hi Jacob,

My guess is that answer (or hint) is here (taken from this link : https://www.phenix-online.org/documentation/faqs/refine.html#targets-and-res...):

*I have ions very close to water molecules/protein atoms, and phenix.refine keeps tring to move them apart. How can I prevent this?*

Use phenix.ready_set or *phenix.metal_coordination* to generate custom bond (and optionally, bond angle) restraints, which will be output to a parameter file ending in ".edits". If you are using the PHENIX GUI, there is a toolbar button for ReadySet in the phenix.refine interface, which will automatically load the output files for use in phenix.refine.

I have not used it myself but I assume that the sigmas for the restraints would be in the *.edits files that you'll generate.

Hope this helps.

Boaz

*Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel E-mail: [email protected] Phone: 972-8-647-2220 Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710 *

------------------------------ *From:* Keller, Jacob [[email protected]] *Sent:* Wednesday, December 30, 2015 4:09 PM *To:* Nigel Moriarty *Cc:* בעז שאנן; [email protected] *Subject:* RE: [phenixbb] Modifications for Ca-binding site

How does one change the ideal bond length/sigma in Phenix?

JPK

*From:* Nigel Moriarty [mailto:[email protected]] *Sent:* Tuesday, December 29, 2015 10:12 AM *To:* Keller, Jacob *Cc:* Boaz Shaanan; [email protected] *Subject:* Re: [phenixbb] Modifications for Ca-binding site

Jacob

Boaz has some sage advice. I recommend not excluding the Ca-O coordination but changing the ideal to better reflect the distances in your files.

PS I'd also like the PDB files, directly, so as not to put noise on the list.

Cheers

Nigel

---

Nigel W. Moriarty Building 33R0349, Physical Biosciences Division Lawrence Berkeley National Laboratory Berkeley, CA 94720-8235 Phone : 510-486-5709 Email : [email protected] Fax : 510-486-5909 Web : CCI.LBL.gov

On Tue, Dec 29, 2015 at 5:49 AM, Keller, Jacob wrote:

Well, I think it's more of a data-vs-geometry battle, with the geometry dragging in the side chains towards the calciums, and the data pulling them out. When I real-space refine them in Coot, some of the Ca-O distances are pretty different from the "ideal" values. Maybe I could just exclude them from xyz refinement in Phenix after making sure they're correct in Coot? I'll give that a shot.

Kol tuv,

Jacob

-----Original Message----- From: Boaz Shaanan [mailto:[email protected]] Sent: Tuesday, December 29, 2015 7:52 AM To: Keller, Jacob; [email protected] Subject: RE: Modifications for Ca-binding site

Hi Jacob,

Sucking-up neighboring residues into the Calcium (or other heavier atoms) density is a known issue. Turning-off the Ca-O or other distance restraints around the Calcium is the last thing you want to do, in my experience. You'd better check those distance restraints and verify that they're compatible with the distances in other (preferably high resolution, of course) structures with calcium binding sites similar to yours and then make sure they're given the appropriate weight during refinement. That's my view but Pavel will give you more advices I'm sure.

Cheers,

Boaz

Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel

E-mail: [email protected] Phone: 972-8-647-2220 Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710

________________________________________ From: [email protected] [ [email protected]] on behalf of Keller, Jacob [ [email protected]] Sent: Tuesday, December 29, 2015 4:44 AM To: [email protected] Subject: [phenixbb] Modifications for Ca-binding site

I am getting a lot of problems with my calcium binding sites--there is clear density for the calciums and the protein side chains, but Phenix keeps sucking the side chains out of their density towards the calciums and also the calciums show some positive difference density, perhaps due to too-high b-factors (or potentially partial occupancy by Na?). I think what might work best is to turn off the Ca-O bond length restraints somehow, but don't see where/how to do that. I tried releasing them from geometrical restraints, but that made things worse, since things went haywire in the sidechains themselves.

Is it possible to make Phenix ignore the Ca-O bond distance restraints?

JPK

******************************************* Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: [email protected] *******************************************

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