Hi Engin,
thank you for all the trouble you went through to make this presentation! The
glycosidic bond in your presentation is indeed correct (alpha1-6).
Not sure how to interpret the terms axial or equatorial; I followed the
definition from here:
https://www.masterorganicchemistry.com/2018/02/19/the-big-damn-post-of-sugar-nomenclature
However, judging from the drawings here
(https://chem.libretexts.org/Bookshelves/Organic_Chemistry/Supplemental_Modules_(Organic_Chemistry)/Alkanes/Properties_of_Alkanes/Cycloalkanes/Rings%3A_cis_trans_and_axial_equatorial_relationships),
yes, the glycosidic bond in my case is equatorial and privateer reports it as
beta1-6.
Its not urgent at all, but if you find a moment, could you please send me the terminal output of
Coot it reports regarding your glycosidic linkages? To do that, start Coot from a terminal and load
your carbohydrate tree. Centre on one of the atoms in your carbohydrate tree, open the "Add
N-linked Glycan" panel in the Glyco module and press "Update for Current Residue".
Coot will print debug info on the terminal.
In the unlike case that terminal output is disabled, you can enable it in
Edit->Preferences->Other->Console->Display state commands in console?->Yes
Could you also send me the coordinates of your carbohydrate tree? It would like
to load it in my Coot version(s) and check the terminal output.
Thanks again!
With best regards,
Markus
On 2024-04-23 19:38, Engin Özkan wrote:
Caution! This message was sent from outside the University of Manitoba.
Hi Markus,
I am using Coot that comes with CCP4 8, which has FUC (alpha-L-Fucose)
with an oxygen on C1 in the axial position. Which is the correct form, I
believe.
Using Coot 0.9.8.93, I get my alpha1-6 and alpha1-3 (insect) Fucoses
inserted correctly. I had trouble understanding your description: You
mean you get Coot to insert a FUC with the glycosidic bond going
equatorial, instead of axial, to Fucose?
This is how it works for me:
https://www.dropbox.com/scl/fi/yjbzstojjivmfpqdlz2os/Screen-Recording-2024-04-23-at-7.32.14-PM.mov?rlkey=zvvk7apd19wk61kt21qyc42l5&dl=0
I'll see what happens when I update to the new Coot version where Paul
has switched the logic.
By the way, privateer tells me I have zero errors and the correct alpha
anomer for Fucose.
Fuc913-a6-GlcNAc911-b-ASN258
WURCS=2.0/2,2,1/[a2122h-1b_1-5_2*NCC/3=O][a1221m-1a_1-5]/1-2/a6-b1
...
PDB Sugar Q Phi Theta Detected type
Cnf <Bfac> Ctx Ok?
---- ------------ ----- ------ ------ -------------------
--- ------ --- -----
test NAG-A-911 0.570 280.48 5.29 beta-D-aldopyranose
4c1 47.88 (n) yes
test FUC-A-913 0.535 92.53 177.32 alpha-L-aldopyranose
1c4 70.76 (n) yes
...
Wrong anomer: 0
Best,
Engin
On 4/18/24 4:35 PM, Markus Meier wrote:
Hi Engin,
hmm, it appears I was using the monomer library from
https://www2.mrc-lmb.cam.ac.uk/personal/pemsley/coot/dependencies/monomers-2023-01-02-23:57:29.tar.gz
(it gets installed automatically with Coot's build-it script).
However, the files in question (FUC.cif, NAG.cif, BMA.cif and MAN.cif)
are identical to current CCP4 8.0.018 version of the library
(installed by CCP4 Update Manager) as well as version ccp4-8.0.018 and
version ccp4-8.0.016 on GitHub
(https://github.com/MonomerLibrary/monomers)
MD5 checksums on these files are
9cfba4ac24f056d648df8227316f788b monomers/f/FUC.cif
958ac5b19a92ceec604d233457944b2a monomers/n/NAG.cif
697dd5242334dd99dad9ce43133c6382 monomers/b/BMA.cif
9d68ff54296ef2f4622005efb4eb080b monomers/m/MAN.cif
If they are different on your machine, please let me know.
I will switch monomer libraries anyway and do some testing.
I also was switching back and forth between my locally compiled coot
(0.9.8.92) and the the CCP4 distributed version (0.9.8.93 EL). I will
check if there is a difference between the two.
Thank you very much for your suggestion!
With best regards,
Markus
On 2024-04-18 13:48, Engin Özkan wrote:
Caution! This message was sent from outside the University of Manitoba.
Hi Markus,
There were reports to the ccp4bb and phenixbb back in 2011 about
something similar. The monomer library files for FUC and/or the linkage
definitions for refmac/phenix were not right. I think. My recollection
is that this was corrected, and I could not reproduce your observation.
I also had private discussions with Garib Murshudov, who were correcting
the glycan monomer problems back in ~2011.
I could find old links to some of the discussions on bulletin board
archives:
https://phenix-online.org/pipermail/phenixbb/2011-November/017958.html
https://www.mail-archive.com/ccp4bb@jiscmail.ac.uk/msg13968.html
I am curious if something got reverted or if you have some ancient
library definitions somewhere. But I hope this gives some ideas.
Best,
Engin
On 4/17/24 3:07 PM, Markus Meier wrote:
Hi Paul,
thank you for implementing the Glyco module in Coot!
I used its "Add N-linked Glycan" function to build an N-linked glycan
which contains L-fucose with an alpha1-6 glycosidic linkage on the
first N-acetyl glucosamine of the tree.
However, after exporting the tree, I checked it with privateer and
unfortunately privateer flags the L-fucose as having the wrong
anomeric linkage - beta 1-6 :-(
Coot reports the same as being in the alpha 1-6 anomeric form.
I then manually built the opposite anomeric form which privateer
happily accepts as being alpha1-6. However, Coot reports it as beta
1-6.
In my understanding, the alpha configuration of a glycosidic linkage
should have the oxygen at the anomeric centre and the -CH3 group at
opposite faces of the sugar ring, so I have to agree with privateer on
this one.
Please check you code.
I have attached the two glycan trees and here is the output of
privateer and Coot for both. Coot and privateer versions are given at
the bottom.
Btw. Neither privateer nor refmac5 can read the mmCIF files that Coot
produces with the "Extract this Tree" functions. All the
_atom_site.label_*_id entries are empty!
N133_tree_privateer_alpha_anomer_FUC.cif:
Privateer:
Detailed validation data
------------------------
PDB Sugar Q Phi Theta Detected type
Cnf <Bfac> Ctx Ok?
---- ------------ ----- ------ ------ -------------------
--- ------ --- -----
N133 NAG-A-459 0.597 63.27 2.85 beta-D-aldopyranose
4c1 28.09 (n) yes
N133 NAG-A-460 0.603 145.39 2.15 beta-D-aldopyranose
4c1 36.34 (n) yes
N133 BMA-A-461 0.550 333.80 4.39 beta-D-aldopyranose
4c1 45.25 (n) yes
N133 MAN-A-462 0.628 6.26 5.03 alpha-D-aldopyranose
4c1 46.22 (n) yes
N133 MAN-A-463 0.628 4.87 5.87 alpha-D-aldopyranose
4c1 50.37 (n) yes
N133 FUC-A-464 0.615 162.25 174.51 alpha-L-aldopyranose
1c4 31.24 (n) yes <-
Partially occupied monosaccharides, if any, are marked with an
asterisk (*)
SUMMARY:
Wrong anomer: 0
Wrong configuration: 0
Unphysical puckering amplitude: 0
In higher-energy conformations: 0
Privateer has identified 0 issues, with 0 of 6 sugars affected.
Coot output:
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.486 A 459 C1 to A 133 ND2
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 133 ASN
returns "pyr-ASN"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 4
#### glyco close: 2.397 A 459 C4 to A 460 C1
#### glyco close: 2.339 A 459 O4 to A 460 C2
#### glyco close: 2.318 A 459 O4 to A 460 O5
#### glyco close: 1.387 A 459 O4 to A 460 C1
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 460 NAG
returns "BETA1-4"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 3
#### glyco close: 2.347 A 460 O4 to A 461 C2
#### glyco close: 2.346 A 460 O4 to A 461 O5
#### glyco close: 1.398 A 460 O4 to A 461 C1
debug:: find_glycosidic_linkage_type() for A 460 NAG,A 461 BMA
returns "BETA1-4"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.407 A 461 O6 to A 462 C1
debug:: find_glycosidic_linkage_type() for A 461 BMA,A 462 MAN
returns "ALPHA1-6"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.403 A 461 O3 to A 463 C1
debug:: find_glycosidic_linkage_type() for A 461 BMA,A 463 MAN
returns "ALPHA1-3"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 2
#### glyco close: 2.385 A 459 O6 to A 464 O5
#### glyco close: 1.399 A 459 O6 to A 464 C1
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 464 FUC
returns "BETA1-6" <-
##########################################################################################
N133_tree_privateer_beta_anomer_FUC.cif:
Privateer:
Detailed validation data
------------------------
PDB Sugar Q Phi Theta Detected type
Cnf <Bfac> Ctx Ok?
---- ------------ ----- ------ ------ -------------------
--- ------ --- -----
N133 NAG-A-459 0.539 93.90 14.28 beta-D-aldopyranose
4c1 17.31 (n) yes
N133 NAG-A-460 0.576 3.57 3.48 beta-D-aldopyranose
4c1 28.29 (n) yes
N133 BMA-A-461 0.534 312.39 9.79 beta-D-aldopyranose
4c1 42.24 (n) yes
N133 MAN-A-462 0.580 219.67 4.19 alpha-D-aldopyranose
4c1 42.02 (n) yes
N133 MAN-A-463 0.563 294.23 4.86 alpha-D-aldopyranose
4c1 47.39 (n) yes
N133 FUC-A-464 0.542 120.60 172.19 beta-L-aldopyranose
1c4 18.87 (n) no <-
Partially occupied monosaccharides, if any, are marked with an
asterisk (*)
SUMMARY:
Wrong anomer: 1
Wrong configuration: 0
Unphysical puckering amplitude: 0
In higher-energy conformations: 0
Privateer has identified 1 issues, with 1 of 6 sugars affected.
Coot:
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.462 A 459 C1 to A 133 ND2
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 133 ASN
returns "pyr-ASN"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 4
#### glyco close: 2.355 A 459 C4 to A 460 C1
#### glyco close: 2.316 A 459 O4 to A 460 O5
#### glyco close: 2.313 A 459 O4 to A 460 C2
#### glyco close: 1.387 A 459 O4 to A 460 C1
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 460 NAG
returns "BETA1-4"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 3
#### glyco close: 2.37 A 460 O4 to A 461 C2
#### glyco close: 2.344 A 460 O4 to A 461 O5
#### glyco close: 1.401 A 460 O4 to A 461 C1
debug:: find_glycosidic_linkage_type() for A 460 NAG,A 461 BMA
returns "BETA1-4"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.408 A 461 O6 to A 462 C1
debug:: find_glycosidic_linkage_type() for A 461 BMA,A 462 MAN
returns "ALPHA1-6"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 3
#### glyco close: 2.39 A 461 C3 to A 463 C1
#### glyco close: 2.389 A 461 O3 to A 463 C2
#### glyco close: 1.398 A 461 O3 to A 463 C1
debug:: find_glycosidic_linkage_type() for A 461 BMA,A 463 MAN
returns "ALPHA1-3"
DEBUG:: find_glycosidic_linkage_type() number of sorted
distances: 1
#### glyco close: 1.414 A 459 O6 to A 464 C1
debug:: find_glycosidic_linkage_type() for A 459 NAG,A 464 FUC
returns "ALPHA1-6" <-
Program versions:
Coot 0.9.8.93 EL (ccp4) on Gentoo Linux 64 bit
Privateer version MKIV : 06/02/2 CCP4 8.0.019 on Gentoo Linux 64 bit
With best regards,
Markus
--
Engin Özkan, Ph.D.
Associate Professor
Dept of Biochemistry and Molecular Biology
University of Chicago
http://ozkan.uchicago.edu
--
Markus Meier, Ph.D.
Research Associate
University of Manitoba
Department of Chemistry
144 Dysart Road
Winnipeg, MB, R3T 2N2, Canada
Phone: +1 204 474 7172
E-mail: markus.me...@umanitoba.ca
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