Hi Marilyn,
if 'some luck' happens not to be the case you may be able to create a
good cif file for the covalent protein-sugar bond with the JLigand
tool (CCP4 suite). that tool allowed me to connect an isoaspartate
residue at both ends with my protein chain so that neither coot nor
refmac blew it up, so I guess it should be able to do the trick for
you, too. if you do not get it to work you can contact me off-list and
I'll send my how-to notes for JLigand.
Bärbel
Zitat von herman.schreu...@sanofi.com:
Hi Marilyn,
I have been struggling with carbohydrates in the pre Glyco module
era, which was *not nice*. However, the following strategy works
reasonably well:
1) Get the XYP monomer with the "get monomer" command.
2) Move the XYP manually in its electron density. This can be
very approximate
3) Do a real space refinement of the XYP monomer alone. This
should work should give a decent fit of the monomer
4) Delete one of the oxygens of the future linkage and use the
"make link" command (extensions-modeling-make link) to link your
monomer to the carbohydrate tree.
For buster, I had some restraints files to link carbohydrates, I do
not have any experience with phenix refine. With some luck, just a
covalent link with reasonably well-defined sugars would be
sufficient. Changing the stereochemistry of a standard carbohydrate
link file may also do the trick.
Good luck!
Herman
Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag
von Yoder, Marilyn
Gesendet: Donnerstag, 5. Juli 2018 16:01
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [EXTERNAL] [ccp4bb] COOT: adding xylose to a plant
glycosylation chain
Hello,
I'm having problems adding XYP (beta-xylose) to my glycosylation
chain in a plant protein.
I have a 1.87 Angstrom map, 4 molecules in the asymmetric unit. I
have two primary N-glycosylation sites, per protein. I am not using
NCS for refinement or model building.
The density for the NAG-(FUC)-NAG is very clear at both sites of all
molecules in the a.u. The BMA (beta mannose after the 2nd NAG) is
clear at one of the glycosylation sites. At this glycosylation
site, I see good density for a MAN at the C3 and C6 sites of BMA. I
can see good density for a xylose at the C2 - it is really quite
clear.
Using COOT, and the Glyco module, set to Hybrid (Plant), I have been
able to add all the NAGs, MANs, and FUCs with a press of a mouse
button. It is a *very nice* function of COOT.
But..... I can't get the XYP added correctly. I select the "add an
XYP-BMA XYP", and it pops the XYP where I want it, but then it
rapidly wiggles away. It is as if it is actively finding an area of
no density (I am not exaggerating). Selecting 'Refine Tree' does
not resolve the problem. I've tried manually moving the xylose
close enough, but this is not working well. It just refines away in
Phenix Refine.
For refinement, I used ReadySet to generate the restraints
(apparently the XYP-BMA XYP does not have restraints in the standard
library - is that correct?)
This is a big enough problem, I am hoping I am just doing something
'big' that is wrong. I'm assuming (perhaps) it is a library
problem. I would appreciate any suggestions.
Software details:
On Windows, WinCoot 0.8.9
On Mac, Phenix version 1.13-29998-000 (Coot is version 0.8.9)
I model build in WinCoot, refine with Phenix on the Mac
Regards,
Marilyn
___________________________________________________________
Marilyn Yoder | 816-235-1986 | SCB 526 | Division of Cell
Biology & Biophysics, University of Missouri-Kansas City
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--
Bärbel Blaum, Ph.D.
Interfakultäres Institut für Biochemie (IFIB)
Hoppe-Seyler-Strasse 4
D-72076 Tübingen
Germany
+49 70 71 29 733 48
Raum 311
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