[phenixbb] Glycan Links/Restraints
Damian Ekiert
dcekiert at scripps.edu
Tue Mar 1 14:09:10 PST 2011
Engin,
Thanks for your help.
Initially, I tried the order you are suggesting for the links ("inner first, outer second"), but Phenix quits with the following output:
Sorry: Traceback (most recent call last):
File "/jcsg/software/phenix/phenix-1.7/phenix-1.7-650/cctbx_project/mmtbx/monomer_library/pdb_interpretation.py", line 972, in apply_mod
mod_mon = self.monomer.apply_mod(mod_mod_id)
File "/jcsg/software/phenix/phenix-1.7/phenix-1.7-650/cctbx_project/mmtbx/monomer_library/cif_types.py", line 387, in apply_mod
result.delete_atom_in_place(mod_atom.atom_id)
File "/jcsg/software/phenix/phenix-1.7/phenix-1.7-650/cctbx_project/mmtbx/monomer_library/cif_types.py", line 304, in delete_atom_in_place
raise RuntimeError("delete_atom_in_place: unknown atom_id: %s" % atom_id)
RuntimeError: delete_atom_in_place: unknown atom_id: O1
apply_mod failure:
pdbres="ASN A 91 "
comp id: ASN
mod id: DEL-O1
This sounded to me like it was trying to delete O1 from my Asn instead of the GlcNAc, which I figured meant that I should reverse the order (i.e., outer residue first, inner residue second). When I did this, phenix completed refinement with no obvious problems in the log file. I initially took this to mean that all was well, until I looked at my coordinates.
As for glycans in the same chain or different chains, I agree with you completely. I actually split them into separate chains to try to get around another apparent problem that was cropping up near the top of my log file:
Chain: "C"
Number of atoms: 5065
Number of conformers: 2
Conformer: "A"
Number of residues, atoms: 327, 5018
Unusual residues: {'MAN%DEL-HO3': 1, 'MAN%DEL-HO4%DEL-O1': 1, 'NAG%DEL-O1': 1, 'NAG%DEL-HO4%DEL-O1': 1}
Classifications: {'undetermined': 4, 'peptide': 323}
Modifications used: {'DEL-HD22': 1, 'DEL-HO3': 1, 'DEL-O1': 3, 'DEL-HO4': 2}
Link IDs: {'PTRANS': 18, None: 4, 'TRANS': 304}
Not linked:
pdbres="NAG C 401 "
pdbres="NAG C 402 "
Not linked:
pdbres="NAG C 402 "
pdbres="BMA C 403 "
Not linked:
pdbres="BMA C 403 "
pdbres="MAN C 404 "
Unresolved non-hydrogen bonds: 1
Unresolved non-hydrogen angles: 2
Unresolved non-hydrogen chiralities: 1
These "Not linked" sections went away when I moved the glycans to separate chains, so I figured that Phenix might only allow one polymer type in a given chain (not sure if this is true or not).
As for the monomer cif files, I don't see an major differences between the cifs from Phenix and those from CCP4 that I have used previously for refinement in Refmac, but I will look into this further.
Best,
Damian
On Feb 28, 2011, at 11:44 PM, Engin Özkan wrote:
> Also, I just remembered that Phenix/monomer library may still not have
> BMA and MAN right. You may have to provide MAN and/or BMA cif files with
> the correct stereochemistries. Monomer library does not necessarily
> follow the standards of the PDB chemical component library (but
> obviously, PDB enforces them).
>
> Finally, you can always check the .geo file that phenix.refine produces,
> and you will find all the applied restraints. That should definitively
> tell you your problem.
>
> Engin
>
> On 2/28/11 10:36 PM, Engin Özkan wrote:
>> Dear Damien,
>>
>> Let me make a guess: You have the BETA1-4 linkages in the wrong order.
>> If you look in the mon_lib_list.cif file, you can find the line where
>> it says:
>> BETA1-4 1 O4 2 C1 single 1.439 0.020
>> which tells me that the residue to be defined (residue_selection_1)
>> ought to be the atom atom connecting with the O4 atom (the inner
>> glycan) and the second residue is the one with the C1 atom (the outer
>> glycan). So try switching residue_selection_1 and _2 for your BETA1-4
>> linkages. I know you are probably thinking BETA1-4 should go 1 to 4,
>> but that is not how it is defined in the monomer library. By the way,
>> your NAG-ASN linkages do look correct, and you have the correct
>> anomers for beta-mannose (BMA) and alpha-mannose (MAN), which is where
>> most people seem to get stuck.
>>
>> One stylistic question I have for you is, why are you naming your
>> protein and glycans with different chain IDs; aren't they actually
>> covalently linked? There apparently is no right way for numbering
>> glycan residues, and everybody seems to be do a different thing...
>>
>> Good luck,
>> Engin
>>
>> On 2/28/11 6:39 PM, Damian Ekiert wrote:
>>> Sorry to bring up an old topic again but I can't find any
>>> documentation of glycan refinement on the Phenix website and previous
>>> threads don't seem to answer my question.
>>>
>>> I am refining a structure with N-linked glycans. I have inserted
>>> several "apply_cif_link" records to my .eff file to define the glycan
>>> topologies (see below) and fed in a cif file for BMA. After fiddling
>>> for quite some time now, I have managed to get around all the error
>>> messages people have described previously to disappear. Yet, 1) none
>>> of my linkages appear to be enforced and 2) even the restraints on
>>> the individual sugars appear to be unenforced (or at least,
>>> insufficiently so). For example, some rings are rendered almost
>>> flat, or flipped into boat or otherwise distorted conformations, and
>>> glycosidic bonds stretch to>2 A.
>>>
>>> No obvious signs of trouble in the log file, running the latest
>>> version (1.7, also tried 1.6.4), but my glycans look terrible
>>> (despite some of the best density I have ever seen! :-) )
>>>
>>> Any suggestions would be appreciated!
>>>
>>> Thanks,
>>>
>>> Damian Ekiert
>>>
>>>
>>>
>>> From my .eff file:
>>>
>>> apply_cif_link {
>>> data_link = NAG-ASN
>>> residue_selection_1 = chain X and resname NAG and resid 401
>>> residue_selection_2 = chain A and resname ASN and resid 91
>>> }
>>> apply_cif_link {
>>> data_link = NAG-ASN
>>> residue_selection_1 = chain Y and resname NAG and resid 401
>>> residue_selection_2 = chain C and resname ASN and resid 91
>>> }
>>> apply_cif_link {
>>> data_link = NAG-ASN
>>> residue_selection_1 = chain Z and resname NAG and resid 401
>>> residue_selection_2 = chain E and resname ASN and resid 91
>>> }
>>> apply_cif_link {
>>> data_link = BETA1-4
>>> residue_selection_1 = chain X and resname NAG and resid 402
>>> residue_selection_2 = chain X and resname NAG and resid 401
>>> }
>>> apply_cif_link {
>>> data_link = BETA1-4
>>> residue_selection_1 = chain Y and resname NAG and resid 402
>>> residue_selection_2 = chain Y and resname NAG and resid 401
>>> }
>>> apply_cif_link {
>>> data_link = BETA1-4
>>> residue_selection_1 = chain Z and resname NAG and resid 402
>>> residue_selection_2 = chain Z and resname NAG and resid 401
>>> }
>>> apply_cif_link {
>>> data_link = BETA1-4
>>> residue_selection_1 = chain Y and resname BMA and resid 403
>>> residue_selection_2 = chain Y and resname NAG and resid 402
>>> }
>>> apply_cif_link {
>>> data_link = ALPHA1-3
>>> residue_selection_1 = chain Y and resname MAN and resid 404
>>> residue_selection_2 = chain Y and resname BMA and resid 403
>>> }
>>> _______________________________________________
>>> phenixbb mailing list
>>> phenixbb at phenix-online.org
>>> http://phenix-online.org/mailman/listinfo/phenixbb
>>
>>
>
>
> --
> Engin Özkan
> Post-doctoral Scholar
> Laboratory of K. Christopher Garcia
> Howard Hughes Medical Institute
> Dept of Molecular and Cellular Physiology
> 279 Campus Drive, Beckman Center B173
> Stanford School of Medicine
> Stanford, CA 94305
> ph: (650)-498-7111
>
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