Hi Ben,

I think you've done the hard part already by building a poly-ala model into your extra density!

If you want to try and sequence it using the density, you can use phenix.replace_side_chains with the keyword "sequence_from_density=True" to do this.  See: https://www.phenix-online.org/documentation/reference/replace_side_chains.html

To use autobuild with this, add an extra line to your sequence file with the sequence of the extra piece (separate it from the previous sequence with a blank line or a ">".)

I would try rebuild_in_place=True to try and optimize the position of your extra chain.  You can specify what residues to work on with something like:

  rebuild_chain_list=A
 rebuild_res_start_list=27
 rebuild_res_end_list=35

This will rebuild chain A27 through A35.

For density modification you might want to use a mask based on your model.  For this you would say:

 mask_from_pdb=my_existing_pdb_file.pdb

Let me know if that doesn't do it!
All the best,
Tom T


Date: Tue, 9 Jan 2018 20:50:41 +0000
From: "Flath, Ben" <bef206@mail.usask.ca>
To: "phenixbb@phenix-online.org" <phenixbb@phenix-online.org>
Subject: [phenixbb] Autobuild in to "extra density"
Message-ID: <52cba352941e48869fc9d5223a97ccaf@Mail06.usask.ca>
Content-Type: text/plain; charset="us-ascii"

Hi All
I have a structure I am building/refining at 2.6 A resolution, there are three NSC molecules in P321 unit cell. Each monomer contains 150 aa and are at least 95% built. Here is where I need some help. There is extra density. Gel and Mass Spec data suggests that there is a smaller MW weight protein/band contained in the crystals which is most likely a fragment of my target protein. Basically I have 3 NCS copies plus a half a molecule that does not appear to have the same tertiary structure (LSQ & SSM do not align well) as the other 3 molecules. I have been able to build several poly-A helices in to this density and R/Rfree are now 0.25/0.31.

My Question is, how do I set up a phenix run (presumably autobuild) So that I can begin to dock my sequence in to these chains and have them extended? Does my sequence input file need to include more than one sequence string? Asking autobuild to find and build 4 NCS groups does not seem to build anything in to this extra density (Also the sigma level for the extra density chains seems to be somewhat less than the well-defined part) I am guessing this is a rebuild_in_place=false job? Can I specify only a certain chain only to be rebuilt? Also are there some special considerations with regard to density modification I should be aware of?

Any help would be much appreciated.

Regards


Benjamin Flath, M.Sc.
Research Assistant, Cygler Lab
University of Saskatchewan
Department of Biochemistry
Room 3D40 HSCI
Box 27, 107 Wiggins Road
Saskatoon, SK
S7N 5E5
Phone: 306 220-7355
email: benjamin.f@usask.ca<mailto:benjamin.f@usask.ca>

--
Thomas C Terwilliger
Laboratory Fellow, Los Alamos National Laboratory
Senior Scientist, New Mexico Consortium
100 Entrada Dr, Los Alamos, NM 87544
Tel: 505-431-0033