Nucleic Acids Research 2007 35(Database issue):287-290; doi:10.1093/nar/gkl907
Nucleic Acids Research, 2007, Vol. 35, No. suppl_1 287-290
© 2006 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
GlycoMapsDB: a database of the accessible conformational space of glycosidic linkages
M. Frank*,
T. Lütteke1 and
C.-W. von der Lieth
German Cancer Research Center, Spectroscopic Department (B090) Molecular Modeling Group, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany
1 Bijvoet Center for Biomolecular Research, Department of Bioorganic Chemistry, Utrecht University Padualaan 8, 3584 CH Utrecht, The Netherlands
*To whom correspondence should be addressed. Tel: +49 6221 424541; Fax: +49 6221 42454; Email: m.frank{at}dkfz.de
Received August 16, 2006. Revised October 11, 2006. Accepted October 12, 2006.
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ABSTRACT
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Conformational energy maps of the glycosidic linkages are a
valuable resource to gain information about preferred conformations
and flexibility of carbohydrates. Here we present GlycoMapsDB,
a new database containing more than 2500 calculated conformational
maps for a variety of di- to pentasaccharide fragments contained
in N- and O-glycans. Oligosaccharides representing branchpoints
of N-glycans are included in the set of fragments, thus the
influence of neighbouring residues is reflected in the conformational
maps. During refinement of new crystal structures, maps contained
in GlycoMapsDB can serve as a valuable resource to check whether
the torsion values of a glycosidic linkage are located in an
‘allowed’ region similar to the Ramachandran plot
analysis for proteins. This might help to improve the structural
quality of the glycan data contained in the Protein Data Bank
(PDB). A link between GlycoMapsDB and the PDB has been established
so that the glycosidic torsions of all glycans contained in
the PDB can be retrieved and compared to calculated data. The
service is available at
www.glycosciences.de/modeling/glycomapsdb/.
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INTRODUCTION
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Ramachandran plots (
1) of protein backbone torsions
/
are frequently
used to validate 3D structures of proteins (
2,
3). The quality
of a protein structure is considered to be ‘good’
when (preferably all) amino acids have
/
values located in ‘allowed’
regions of the plot. For carbohydrates the glycosidic torsions
/
(
) are the main determinants of the 3D structure and it is
straightforward to validate the quality of an experimentally
determined carbohydrate structure in a similar way to protein
structures. In contrast to proteins, however, the ‘allowed’
regions on a conformational map for a given glycosidic linkage
not only depend on the linked monosaccharide types, but also
on the linkage type and—a feature completely different
from proteins—the degree of branching of the glycan. The
number of available high quality crystal structures of carbohydrates
is too limited to serve as a basis to determine the ‘allowed’
regions for all linkage types so the accessible conformational
space of carbohydrate linkages has therefore to be estimated
using computational methods. Consequently, over the last 30
years a considerable effort has been put into the development
of force fields that are able to predict accurately the local
minima and flexibility of glycosidic torsions (
4). The force
field most frequently used to calculate conformational maps
of glycosidic linkages is MM3 (
5).
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CALCULATION OF CONFORMATIONAL MAPS
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A variety of methods exist to calculate the energy of a carbohydrate
as a function of the glycosidic torsions
/
(or
/
/
for 1–6
linkages). Traditionally systematic search methods are applied
to disaccharides (
6). A ‘relaxed’ map is obtained
by systematically changing
and
in small intervals (normally
10°) and minimizing all degrees of freedom while restraining
/
using an external force. Relaxed conformational maps depend
on the orientation of the exocyclic torsions in the starting
conformation of the carbohydrate and therefore the calculation
of an ‘adiabatic’ map is advisable. To generate
a fully adiabatic map the calculation of 3
10 relaxed maps would
be required for a simple disaccharide (
7). In routine calculations
the computational cost is reduced by taking into account only
gg and
gt (or
gt and
tg for monosaccharides that have the OH4
group in an axial orientation) conformations for the hydroxymethyl
groups and a
clock- and
anticlockwise orientation for the hydroxyl
groups. This reduces the number of relaxed maps to be calculated
to eight resulting in a total of 10 368 conformations to minimize
for each ‘pseudo’ adiabatic map. It is obvious that
this approach is limited to disaccharides since the number of
conformations to be minimized for a trisaccharide would already
be
3
x 10
6 (assuming the second linkage is searched in intervals
of 30°), far too many for a large-scale project where thousands
of maps need to be calculated.
High temperature molecular dynamics simulation is a robust and efficient method to explore the accessible conformational space of carbohydrates (8). Conformational free energy maps can be derived from population analysis by applying the Boltzmann equation. This approach has several advantages compared to the systematic search methods:
- it is directly applicable to branched oligosaccharides, so that the same method can be used for disaccharides and larger oligosaccharides,
- the low energy conformational space only is explored and no computational time is wasted to calculate unrealistic high energy conformations,
- the required computational cost increases therefore only moderately with the number of atoms of the oligosaccharide,
- the data-flow can be easily optimised in such a way that the conformational maps are generated automatically and only minimal human interaction is required.
It was therefore decided that free energy maps derived from MD simulations were a good first set of conformational maps with which to populate the GlycoMaps database. An in-house library of carbohydrate fragments (up to pentasaccharides) derived from structures described in the CARBBANK (9) and built using the SWEET-II program (10) served as input. The MM3 force field as implemented in the TINKER suite (dasher.wustl.edu/tinker/) [for a comparision with the original MM3 implementation see (11)] was used to calculate the trajectories at 1000 K. The length of the MD simulation was 10 ns for disaccharides and 30 ns for larger oligosaccharides. The carbohydrate rings were restrained to a chair conformation. The Conformational Analysis Tools (CAT) software (www.md-simulations.de/CAT/) was used for data processing and analysis.
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ACCESSING AND ANALYSING CONFORMATIONAL MAPS
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Conformational maps of carbohydrate linkages can be retrieved
from the database by entering the disaccharide fragment in IUPAC
form into the search database web interface of GlycoMapsDB (
Figure 1).
Wildcards are supported, which allows searching, e.g. Glcp and
Glcp2NAc residues simultaneously. Maps found in the database
are displayed as a list, with each entry showing a preview picture
and the full structure of the carbohydrate for which the map
has been calculated in extended IUPAC form (
Figure 2). Difference
maps can be calculated e.g. to evaluate the influence of branching
on the accessible conformational space of a linkage (
Figure 3).
Individual maps can be explored in more detail by clicking on
the preview picture. A 3D structure of the carbohydrate—generated
by Sweet II (
10)—can be displayed using JMol (
jmol.sourceforge.net).
If experimental data for the disaccharide fragment is available
in the Protein Data Bank (PDB) (
12), a link is displayed, which
leads to a page where
/
torsion values retrieved from PDB data
[using the GlyTorsion tool (
13)] are overlaid onto the calculated
conformational map (
Figure 4). Some statistical data regarding
the fit of the experimental data to the calculated data is also
displayed. The PDB structures evaluated can be accessed following
the ‘show details’ link. If further information
such as literature references or NMR data for the carbohydrate
is available in GlycosciencesDB (
14) a link to the corresponding
entry in that database is displayed.
View larger version (55K):
[in this window]
[in a new window]
[Download PowerPoint slide]
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Figure 2 Example of a search result. The complete carbohydrate structure used for the calculation of the conformational map is displayed in IUPAC extended nomenclature. The disaccharide fragment that defines the glycosidic linkage is highlighted. The method used to calculate the maps is shown as well as a preview picture of the map that serves as an active link to a more detailed display of the database entry.
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View larger version (60K):
[in this window]
[in a new window]
[Download PowerPoint slide]
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Figure 3 Difference maps can be calculated to compare conformational maps. In the example shown the conformational maps of the disaccharide fragment ß-D-GlcpNAc-(1-4)- -D-Manp either as part of a highly branched oligosaccharide (b) or as part of a linear chain (c) are compared. The reduction of accessible conformational space caused by neighbouring residues in the branched oligosaccharide can clearly be seen in the difference map (a, deep red areas represent regions where the neighbouring residue causes a strong energy penalty mainly by steric conflicts).
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View larger version (54K):
[in this window]
[in a new window]
[Download PowerPoint slide]
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Figure 4 Comparison of calculated conformational maps with glycosidic torsion values derived from the PDB using the GlyTorsion tool (13). The PDB database entries can be explored using the ‘show details’ link. If available, literature references and NMR data for the carbohydrate can be retrieved from GlycosciencesDB.
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IMPLEMENTATION
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GlycoMapsDB is running on a Linux PC with Apache web server
software. Interaction with the user is mediated through PHP
interfaces. The datasets are stored in a mySQL database. Visualization
of carbohydrate structures is performed using the java applet
Jmol or the plugin Chime (
www.mdlchime.com). Diagrams and plots
are generated in scalable vector graphics (SVG) format. For
browsers that cannot display SVG files, they are converted to
graphics interchange format (GIF) files using ImageMagick. The
service is hosted at and maintained by the German Cancer Research
Centre in Heidelberg, Germany. The database can be accessed
online at
www.glycosciences.de/modeling/glycomapsdb/.
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DISCUSSION AND OUTLOOK
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Currently, the GlycoMapsDB contains
2500 conformational maps
of carbohydrate fragments originally described in the CARBBANK.
Conformational maps for most fragments found in glycoproteins
are available in the database. The direct crosslink between
calculated maps and PDB data opens a very efficient route to
crosscheck the quality of experimental structures as well as
the quality of the maps. In general the amount of available
high quality experimental data currently available for carbohydrates
in the PDB database is rather limited compared to that for proteins
and for some linkages there is no experimental data available
at all, so there is a clear lack of experimental reference data.
In this respect, the conformational maps contained in GlycoMapsDB
might help crystallographers to crosscheck their data before
submission similar to the Ramachandran plot analysis for proteins.
For this purpose, maps from the GlycoMapsDB are also accessed
by the
carp (Carbohydrate Ramachandran Plot) software (
13),
where users can upload a structure in PDB file format and retrieve
plots comparing the torsions present in the structure with the
conformational maps.
GlycoMapsDB indirectly offers an interface for ‘data mining’ in the PDB database, e.g. to find carbohydrate entries with unusual glycosidic torsion values. For ß-D-GlcpNAc-(1-4)-ß-D-GlcpNAc, a frequent fragment contained in N-glycans, the agreement between experimental and calculated data is remarkably good (Figure 4). More than 80% of the crystal structures have values in low energy areas of the conformational map. But there are also some outliers that are located in ‘not allowed’ areas of the maps. Of special interest are also those structures that have or values of 180° (anti conformation). The ‘show details’ link would help to find the corresponding entries in the PDB.
In the near future we will provide also conformational maps calculated with force fields other than MM3, so that the influence of different parameter sets can be investigated. An upload feature for maps will be added so that users can upload and compare their own calculated maps with the maps stored in the database. In addition, a functionality that will allow users to calculate maps not already contained in the database is planned.
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ACKNOWLEDGEMENTS
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The development of GlycMapsDB as part of the GLYCOSCIENCES.de
portal at the DKFZ was supported by a Research Grant from the
German Research Foundation (DFG BIB 46 HDdkz 01-01) within the
digital library program as well as the president fond of the
Helmholtz society. Funding to pay the Open Access publication
charges for this article was provided by the DKFZ.
Conflict of interest statement. None declared.
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Footnotes
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Present address:
T. Lütteke, Massachusetts Institute of Technology, Biological Engineering, 77 Massachusetts Avenue, 16-719 Cambridge, MA 0213, USA
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