DTT mediated disulfide crosslink across a crystallographic two-fold!
Hi Phenix Users, I have found a beautifully ordered DTT molecule stretching across a crystallographic two-fold and linking two symmetry related cysteine side chains. Since part of this molecule sits on across the two-fold (special position) I will specify the occupancy as 0.5. I am hoping to get some information on how easy (or how difficult) it is to specify this arrangement in Phenix for refinement since it is covalent and sitting across a crystallographic two-fold (and presumably stabilizing the lattice). Will ReadySet build the appropriate CIF file if I build the DTT into the density and give it a 0.5 occupancy? Joe ___________________________________________________________ Joseph P. Noel, Ph.D. Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: [email protected] Web Site (Salk): http://www.salk.edu/faculty/faculty_details.php?id=37 Web Site (HHMI): http://hhmi.org/research/investigators/noel.html ___________________________________________________________
Hi Joe, I think all you need is summarized here: http://www.phenix-online.org/pipermail/phenixbb/2009-December/003070.html Please let us know if you still have any questions. ReadySet creates a CIF file wich has nothing to do with the occupancies, symmetry etc... It just defines the geometry of your DTT. So, create a CIF file for DTT (if you really need it - I thought it is standard thing and might be already in the library) and run the refinement following the tips in the link above. Pavel. On 8/3/10 5:54 PM, Joseph Noel wrote:
Hi Phenix Users,
I have found a beautifully ordered DTT molecule stretching across a crystallographic two-fold and linking two symmetry related cysteine side chains. Since part of this molecule sits on across the two-fold (special position) I will specify the occupancy as 0.5. I am hoping to get some information on how easy (or how difficult) it is to specify this arrangement in Phenix for refinement since it is covalent and sitting across a crystallographic two-fold (and presumably stabilizing the lattice). Will ReadySet build the appropriate CIF file if I build the DTT into the density and give it a 0.5 occupancy?
Joe ___________________________________________________________ Joseph P. Noel, Ph.D. Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA
Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: [email protected] mailto:[email protected]
Web Site (Salk): http://www.salk.edu/faculty/faculty_details.php?id=37 Web Site (HHMI): http://hhmi.org/research/investigators/noel.html ___________________________________________________________
_______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb
Thanks so much Pavel for the very rapid response! One more thing, it is a disulfide linked DTT so do I do anything special to ensure that all the sulfur atoms (4 in total - two in one unit cell - one each from the cys and dtt and two in the other unit cell) stay in close enough contact for a covalent bond and not move due to non-bonded contact distance restraints? ___________________________________________________________ Joseph P. Noel, Ph.D. Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: [email protected] Web Site (Salk): http://www.salk.edu/faculty/faculty_details.php?id=37 Web Site (HHMI): http://hhmi.org/research/investigators/noel.html ___________________________________________________________ On Aug 3, 2010, at 6:05 PM, Pavel Afonine wrote:
Hi Joe,
I think all you need is summarized here:
http://www.phenix-online.org/pipermail/phenixbb/2009-December/003070.html
Please let us know if you still have any questions.
ReadySet creates a CIF file wich has nothing to do with the occupancies, symmetry etc... It just defines the geometry of your DTT. So, create a CIF file for DTT (if you really need it - I thought it is standard thing and might be already in the library) and run the refinement following the tips in the link above.
Pavel.
On 8/3/10 5:54 PM, Joseph Noel wrote:
Hi Phenix Users,
I have found a beautifully ordered DTT molecule stretching across a crystallographic two-fold and linking two symmetry related cysteine side chains. Since part of this molecule sits on across the two-fold (special position) I will specify the occupancy as 0.5. I am hoping to get some information on how easy (or how difficult) it is to specify this arrangement in Phenix for refinement since it is covalent and sitting across a crystallographic two-fold (and presumably stabilizing the lattice). Will ReadySet build the appropriate CIF file if I build the DTT into the density and give it a 0.5 occupancy?
Joe ___________________________________________________________ Joseph P. Noel, Ph.D. Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA
Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: [email protected]
Web Site (Salk): http://www.salk.edu/faculty/faculty_details.php?id=37 Web Site (HHMI): http://hhmi.org/research/investigators/noel.html ___________________________________________________________
_______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb
Hi Joe,
Thanks so much Pavel for the very rapid response! One more thing, it is a disulfide linked DTT so do I do anything special to ensure that all the sulfur atoms (4 in total - two in one unit cell - one each from the cys and dtt and two in the other unit cell) stay in close enough contact for a covalent bond and not move due to non-bonded contact distance restraints?
phenix.refine shows a summary of the disulfides like this: Number of disulfides: simple=1, symmetry=1 Simple disulfide: pdb=" SG CYS A 2 " - pdb=" SG CYS A 14 " distance=2.09 Symmetry disulfide: pdb=" SG BCYS A 133 " - pdb=" SG BCYS A 133 " distance=2.12 -x,y,-z The program should find all disulfides automatically, using the disulfide_distance_cutoff=3 parameter. Let us know if the output doesn't look right. Ralf
participants (3)
-
Joseph Noel -
Pavel Afonine -
Ralf W. Grosse-Kunstleve