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Dear Clemens,
<div class=""><br class="">
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<div class="">I was a bit sidetracked the last days, but finally today I could test your suggestions. It worked as a charm, both chains. Terrific!</div>
<div class=""><br class="">
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<div class="">Thank you very much! </div>
<div class=""> </div>
<div class="">Please accept my wishes of a Merry Christmas, and a fantastic 2023 - to you and to all the team.</div>
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<div class="">Best,</div>
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<div class="">
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<div>---------------------------------------------------<br class="">
Carlos KIKUTI, PhD<br class="">
UMR144 - CNRS - Institut Curie</div>
<div>Pavillon Trouillet Rossignol<span class="Apple-tab-span" style="white-space: pre;">
</span><br class="">
26 Rue d’Ulm - 75005 Paris, France<br class="">
+33 6 82 87 62 76<br class="">
<a href="mailto:carlos.kikuti@curie.fr" class="">carlos.kikuti@curie.fr</a><span class="Apple-tab-span" style="white-space: pre;">
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<div><br class="">
<blockquote type="cite" class="">
<div class="">Le 12 déc. 2022 à 17:45, Clemens Vonrhein <<a href="mailto:vonrhein@globalphasing.com" class="">vonrhein@globalphasing.com</a>> a écrit :</div>
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<div class="">
<div class="">Dear Carlos,<br class="">
<br class="">
On Wed, Dec 07, 2022 at 09:02:23PM +0000, Kikuti Carlos wrote:<br class="">
<blockquote type="cite" class="">I’m refining a structure at 2.5 angstroms (2.2 with Staraniso)<br class="">
containing nucleotide + hexacoordinated Mg2+, but some of the side<br class="">
chains in the active site seem to be pushed away from the ion,<br class="">
</blockquote>
<br class="">
Yes, if we don't have particular distance/bond restraints between two<br class="">
atoms, a normal non-bonded interaction kicks in (avoiding atoms<br class="">
getting too close). One can avoid the latter by using so-called<br class="">
EXCLUDE cards, e.g.<br class="">
<br class="">
EXCLUDE H|604 H|723<br class="">
<br class="">
(involving all atoms from residues 604 and 723 in the H<br class="">
chain). However, a proper restraint is the better idea, yes.<br class="">
<br class="">
<blockquote type="cite" class="">and I am not sure the geometry of the phosphate moiety-Mg2+ bonds is<br class="">
correct . In this structure, Mg2+ is surrounded by 2 water<br class="">
molecules, 2 phosphate moieties from the nucleotide, two different<br class="">
residues from the myosin. Bond lengths are between 1.8 and 2.3<br class="">
angstroms.<br class="">
</blockquote>
<br class="">
Sounds sensible: the angles are probably close to the canonical 90/180<br class="">
degree as well?<br class="">
<br class="">
<blockquote type="cite" class="">This is a very common scenario in the lab, so your answer<br class="">
will potentially help improving many other future structures,<br class="">
especially the ones in lower resolution, and I’ve got a bunch of<br class="">
those.<br class="">
<br class="">
What I’ve tried already, using the Nov/22 version of Buster:<br class="">
<br class="">
LINK records -> well… I thought CCP4/PDB were recommending this as<br class="">
the way to deal with all kinds of bonds (am I right?), but<br class="">
Buster/TNT/Gelly prefer to use LINK only for covalent bonds. My LINK<br class="">
records are filtered out from the beginning. Buster tells me to «<br class="">
manually describe coordination » , which is the main reason of this<br class="">
e-mail.<br class="">
</blockquote>
<br class="">
Ah LINK records and the different usages of those ;-)<br class="">
<br class="">
According to the PDB documentation at<br class="">
<br class="">
<a href="http://www.wwpdb.org/documentation/file-format-content/format33/sect6.html#LINK" class="">http://www.wwpdb.org/documentation/file-format-content/format33/sect6.html#LINK</a><br class="">
<br class="">
we have<br class="">
<br class="">
The LINK records specify connectivity between residues that is not<br class="">
implied by the primary structure. Connectivity is expressed in terms<br class="">
of the atom names. They also include the distance associated with<br class="">
the each linkage following the symmetry operations at the end of<br class="">
each record.<br class="">
<br class="">
Details<br class="">
<br class="">
* The atoms involved in bonds between HET groups or between a HET<br class="">
group and standard residue are listed.<br class="">
<br class="">
So these are intended mostly as descriptions of existing bonds not<br class="">
implicit in standard residue ordering (furthermore, the distance given<br class="">
is not a restraint distance but the distance observed on the actual<br class="">
ATOM/HETATM records).<br class="">
<br class="">
At some point, REFMAC introduced non-standard LINKR records with a<br class="">
connectivity type specification at the end (which then pointed into an<br class="">
actual restraint dictionary). But being non-standard, there was no way<br class="">
of relying on this really.<br class="">
<br class="">
<blockquote type="cite" class="">(This doesn’t look like the classic formatting/spacing issue as the<br class="">
error message would be different; and I also tried to invert the<br class="">
order of the ligands in the records, no success)<br class="">
</blockquote>
<br class="">
Yes, BUSTER doesn't use these LINK records to establish some kind of<br class="">
connectivity - also because in the old days (still?) there was a lot<br class="">
of random LINK record generation happening at deposition time without<br class="">
manual confirmation and therefore very unreliable.<br class="">
<br class="">
<blockquote type="cite" class="">"good”(?) and old CONECT records -> nah… Buster just ignores them,<br class="">
the output doesn’t have any. I can’t restrain the bond lengths with<br class="">
this anyways.<br class="">
</blockquote>
<br class="">
Correct.<br class="">
<br class="">
<blockquote type="cite" class="">"-Gelly file.dat" with NOTE_BUSTER_DISTANCE… -> from the log it<br class="">
seems this won’t work for HETATM, only for ATOM lines. Maybe there<br class="">
is something I should do in the .seq file?<br class="">
</blockquote>
<br class="">
That's the way to get started, yes. You /could/ add it to the TNT<br class="">
sequence file, but a cleaner way would be a separate text file with<br class="">
those so-called Gelly commands ("utility restraints"):<br class="">
<br class="">
<a href="https://www.globalphasing.com/buster/manual/gelly/manual/gelly5.html" class="">https://www.globalphasing.com/buster/manual/gelly/manual/gelly5.html</a><br class="">
<br class="">
What you want is e.g. (7DY1 as a random example, MG D604)<br class="">
<br class="">
NOTE six coordinating atoms around MG MG D 604 <br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|750:O<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|750:O D|604:MG D|819:O<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|750:O D|604:MG D|295:OG1<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|750:O D|604:MG D|602:O3G<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|750:O D|604:MG D|602:O1B<br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|754:O<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|754:O D|604:MG D|819:O<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|754:O D|604:MG D|295:OG1<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|754:O D|604:MG D|602:O3G<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|754:O D|604:MG D|602:O1B<br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|819:O<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|819:O D|604:MG D|295:OG1<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|819:O D|604:MG D|602:O3G<br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|295:OG1<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|295:OG1 D|604:MG D|602:O1B<br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|602:O3G<br class="">
NOTE BUSTER_UTILANGLE 90.00 1.60 D|602:O3G D|604:MG D|602:O1B<br class="">
NOTE BUSTER_DISTANCE =2.12 0.05 D|604:MG D|602:O1B<br class="">
<br class="">
This defines those 6 distance (here: 2.12A with esd of 0.05) and all<br class="">
90 degree angles (esd = 1.6).<br class="">
<br class="">
<blockquote type="cite" class="">(I’ve found an old presentation from Clemens that shows a<br class="">
hexacoordinated Mg2+. That made me salivate, but the protocol is not<br class="">
described)<br class="">
</blockquote>
<br class="">
We always wanted to push the jiffy/script used at the time into a<br class="">
normal BUSTER distribution, since the generation of the above<br class="">
restraint example is tedious for anything larger. Maybe it is time to<br class="">
get back to this and finally make it available ...<br class="">
<br class="">
In the meantime and if you have only a few Mg ions to handle: define<br class="">
the six distances (you might want to play a bit with value/esd) and<br class="">
all 12 right angles as above. Afterwards, add this text file to your<br class="">
command-line as<br class="">
<br class="">
refine -Gelly Mg-restraints.txt ...<br class="">
<br class="">
or similar.<br class="">
<br class="">
Cheers<br class="">
<br class="">
Clemens, Gerard & Andrew<br class="">
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