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Computer-Assisted Carbohydrate Structural Studies and Drug Discovery
Stockholm University, Faculty of Science, Department of Organic Chemistry. (Göran Widmalm)
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Carbohydrates are abundant in nature and have functions ranging from energy storage to acting as structural components. Analysis of carbohydrate structures is important and can be used for, for instance, clinical diagnosis of diseases as well as in bacterial studies. The complexity of glycans makes it difficult to determine their structures. NMR spectroscopy is an advanced method that can be used to examine carbohydrates at the atomic level, but full assignments of the signals require much work. Reliable automation of this process would be of great help. Herein studies of Escherichia coli O-antigen polysaccharides are presented, both a structure determination by NMR and also research on glycosyltransferases which assemble the polysaccharides. The computer program CASPER has been improved to assist in carbohydrate studies and in the long run make it possible to automatically determine structures based only on NMR data.

Detailed computer studies of glycans can shed light on their interactions with proteins and help find inhibitors to prevent unwanted binding. The WaaG glycosyltransferase is important for the formation of E. coli lipopolysaccharides. Molecular docking analyses of structures confirmed to bind this enzyme have provided information on how inhibitors could be composed. Noroviruses cause gastroenteritis, such as the winter vomiting disease, after binding human histo-blood group antigens. In one of the projects, fragment-based docking, followed by molecular dynamics simulations and binding free energy calculations, was used to find competitive binders to the P domain of the capsid of the norovirus VA387. These novel structures have high affinity and are a very good starting point for developing drugs against noroviruses. The protein targets in these two projects are carbohydrate binding, but the techniques are general and can be applied to other research projects.

Place, publisher, year, edition, pages
Stockholm: Department of Organic Chemistry, Stockholm University , 2011. , 78 p.
Keyword [en]
Carbohydrates, molecular docking, molecular dynamics simulation, fragment-based virtual screening, NMR spectroscopy, computer-aided drug design, computer-aided structure elucidation, glycosyltransferases, Escherichia coli
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
URN: urn:nbn:se:su:diva-56411ISBN: 978-91-7447-245-5OAI: oai:DiVA.org:su-56411DiVA: diva2:411668
Public defence
2011-05-25, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: Submitted. Paper 5: Manuscript. Paper 6. Manuscript.Available from: 2011-05-03 Created: 2011-04-15 Last updated: 2012-07-03Bibliographically approved
List of papers
1. Structural studies of the O-antigenic polysaccharides from the enteroaggregative Escherichia coli strain 94/D4 and the international type strain Escherichia coli O82
Open this publication in new window or tab >>Structural studies of the O-antigenic polysaccharides from the enteroaggregative Escherichia coli strain 94/D4 and the international type strain Escherichia coli O82
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2009 (English)In: Carbohydrate Research, ISSN 0008-6215, E-ISSN 1873-426X, Vol. 344, no 18, 2528-2532 p.Article in journal (Refereed) Published
Abstract [en]

The structure of the O-antigen polysaccharides (PS) from the enteroaggregative Escherichia coli strain 94/D4 and the international type strain E. coli O82 have been determined. Component analysis and 1H, 13C, and 31P NMR spectroscopy experiments were employed to elucidate the structure. Inter-residue correlations were determined by 1H, 13C-heteronuclear multiple-bond correlation, and 1H, 1H-NOESY experiments. d-GroA as a substituent is linked via its O-2 in a phosphodiester-linkage to O-6 of the α-d-Glcp residue. The PS is composed of tetrasaccharide repeating units with the following structure:

→4)-α-d-Glcp6-(P-2-d-GroA)-(1→4)-β-d-Galp-(1→4)-β-d-Glcp-(1→3)-β-d-GlcpNAc-(1→

Cross-peaks of low intensity from an α-d-Glcp residue were present in the NMR spectra and spectral analysis indicates that they originate from the terminal residue of the polysaccharide. Consequently, the biological repeating unit has a 3-substituted N-acetyl-d-glucosamine residue at its reducing end. Enzyme immunoassay using specific anti-E. coli O82 rabbit sera showed identical reactivity to the LPS of the two strains, in agreement with the structural analysis of their O-antigen polysaccharides.

Keyword
Escherichia coli, Lipopolysaccharide, NMR, Biological repeating unit, Glyceric acid
National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-31498 (URN)10.1016/j.carres.2009.09.033 (DOI)000272860500014 ()
Available from: 2009-11-18 Created: 2009-11-18 Last updated: 2011-04-19Bibliographically approved
2. Glycosyltransferase functions of E. coli O-antigens
Open this publication in new window or tab >>Glycosyltransferase functions of E. coli O-antigens
2010 (English)In: Glycobiology, ISSN 0959-6658, E-ISSN 1460-2423, Vol. 20, no 3, 366-368 p.Article in journal (Refereed) Published
Abstract [en]

ECODAB (the E. coli O-antigen database) has been expanded to include information about glycosyltransferases (GTs) involved in the assembly of the O-antigen polysaccharide. Similarity searches have been performed to be able to determine GT functions that have not been reported prior to this work. In addition to suggesting the function of 179 GTs, the approach leads to the prediction of part of the O-antigen structures of a number of serogroups. The procedure suggests a novel way of combining genetic information with experimental techniques in structural analysis of oligo- and polysaccharides.

Keyword
database, Escherichia coli, glycosyltransferases, O-antigen, polysaccharides
National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-37218 (URN)10.1093/glycob/cwp185 (DOI)000274341300010 ()
Available from: 2010-02-17 Created: 2010-02-17 Last updated: 2011-11-23Bibliographically approved
3. Structural analysis of glycans by NMR chemical shift prediction
Open this publication in new window or tab >>Structural analysis of glycans by NMR chemical shift prediction
2011 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 83, no 5, 1514-1517 p.Article in journal (Refereed) Published
Abstract [en]

Structural determination of N- and O-linked glycans as well as polysaccharides is hampered by the limited spectral dispersion. The computerized approach CASPER, an acronym for computer assisted spectrum evaluation of regular polysaccharides, uses liquid state NMR data to elucidate carbohydrate structure based on agreement with predicted 1H and 13C chemical shifts. We here demonstrate developments based on multiple through-bond J-based correlations that significantly enhance the credence to the sequence connectivities proposed in the analysis exemplified by an oligosaccharide and a bacterial polysaccharide. The approach is also suitable for predicting 1H and 13C NMR chemical shifts of synthesized oligosaccharides and glycoconjugates, thereby corroborating a proposed structure.

National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-55136 (URN)10.1021/ac1032534 (DOI)000287685800006 ()
Funder
Swedish Research CouncilKnut and Alice Wallenberg Foundation
Available from: 2011-03-02 Created: 2011-03-02 Last updated: 2012-01-19Bibliographically approved
4. Automatic structure determination of regular polysaccharides based solely on NMR spectroscopy
Open this publication in new window or tab >>Automatic structure determination of regular polysaccharides based solely on NMR spectroscopy
2011 (English)In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 12, no 11, 3851-3855 p.Article in journal (Refereed) Published
Abstract [en]

The structural analysis of polysaccharides requires that the sugar components and their absolute configurations are determined. We here show that this can be performed based on NMR spectroscopy by utilizing butanolysis with (+)- and (-)-2-butanol that gives the corresponding 2-butyl glycosides with characteristic 1H and 13C NMR chemical shifts. The subsequent computer-assisted structural determination by CASPER can then be based solely on NMR data in a fully automatic way as shown and implemented herein. The method is additionally advantageous in that reference data only have to be prepared once and from a user's point of view only the unknown sample has to be derivatized for use in CASPER.

National Category
Organic Chemistry Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:su:diva-70868 (URN)10.1021/bm201169y (DOI)000296830600003 ()
Funder
Swedish Research CouncilKnut and Alice Wallenberg Foundation
Note

authorCount :3

Available from: 2012-01-25 Created: 2012-01-24 Last updated: 2013-09-19Bibliographically approved
5. Exploration of the molecular recognition and dynamic properties of WaaG glycosyltransferase
Open this publication in new window or tab >>Exploration of the molecular recognition and dynamic properties of WaaG glycosyltransferase
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(English)Manuscript (preprint) (Other academic)
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-30118 (URN)
Available from: 2009-10-04 Created: 2009-10-04 Last updated: 2011-04-19Bibliographically approved
6. Fragment-based design of norovirus inhibitors by molecular docking and LIE binding free energy calculations
Open this publication in new window or tab >>Fragment-based design of norovirus inhibitors by molecular docking and LIE binding free energy calculations
(English)Manuscript (preprint) (Other academic)
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-56463 (URN)
Available from: 2011-04-18 Created: 2011-04-18 Last updated: 2011-04-19Bibliographically approved

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