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Structural studies of the O‐antigen polysaccharide from Escherichia coli TD2158 having O18 serogroup specificity and aspects of its interaction with the tailspike endoglycosidase of the infecting bacteriophage HK620
Stockholm University, Faculty of Science, Department of Organic Chemistry. (Göran Widmalm)
Stockholm University, Faculty of Science, Department of Organic Chemistry.
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2012 (English)In: Carbohydrate Research, ISSN 0008-6215, E-ISSN 1873-426X, Vol. 357, 118-125 p.Article in journal (Refereed) Published
Abstract [en]

We have analyzed the O-antigen polysaccharide of the previously uncharacterized Escherichia coli strain TD2158 which is a host of bacteriophage HK620. This bacteriophage recognizes and cleaves the polysaccharide with its tailspike protein (TSP). The polysaccharide preparation as well as oligosaccharides obtained from HK620TSP endoglycosidase digests were analyzed with NMR spectroscopy. Additionally, sugar analysis was performed on the O-antigen polysaccharide and MALDI-TOF MS was used in oligosaccharide analysis. The present study revealed a heterogeneous polysaccharide with a hexasaccharide repeating unit of the following structure: α-D-Glcp-(1→6|) →2)-α-L-Rhap-91→6)-α-D-Glcp-(1→4)-α-D-Ga|lp-(1→3)-α-D-GlcpNAc-(1→ β-D-Glcp/β-D-GlcpNAc-(1→3) A repeating unit with a D-GlcNAc substitution of D-Gal has been described earlier as characteristic for serogroup O18A1. Accordingly, we termed repeating units with D-Glc substitution at D-Gal as O18A2. NMR analyses of the polysaccharide confirmed that O18A1- and O18A2-type repeats were present in a 1:1 ratio. However, HK620TSP preferentially bound the D-GlcNAc-substituted O18A1-type repeating units in its high affinity binding pocket with a dissociation constant of 140 μM and disfavored the O18A2-type having a β-D-Glcp-(1→3)-linked group. As a result, in hexasaccharide preparations, O18A1 and O18A2 repeats were present in a 9:1 ratio stressing the clear preference of O18A1-type repeats to be cleaved by HK620TSP.

Place, publisher, year, edition, pages
Elsevier, 2012. Vol. 357, 118-125 p.
Keyword [en]
Escherichia coli, Tailspike, Endoglycosidase, Lipopolysaccharide, NMR, Mass spectrometry
National Category
Organic Chemistry
Research subject
Organic Chemistry
URN: urn:nbn:se:su:diva-75048DOI: 10.1016/j.carres.2012.05.022OAI: diva2:513855
Swedish Research CouncilKnut and Alice Wallenberg Foundation
Available from: 2012-04-03 Created: 2012-04-03 Last updated: 2013-01-09Bibliographically approved
In thesis
1. Structural and Conformational Studies of Oligo- and Polysaccharides
Open this publication in new window or tab >>Structural and Conformational Studies of Oligo- and Polysaccharides
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The focus of this thesis is to examine the structural properties of polysaccharides produced by bacteria, as well as the dynamic and conformational behavior of a synthetically derived oligosaccharide.

The primary structures of the O-polysaccharide repeating units of four different Escherichia coli (E. coli) strains, namely O175, O177, O103 and TD2158, as well as the first report of a capsular polysaccharide produced by lactic acid bacteria Leuconostoc mesenteroides ssp. cremoris PIA2 are reported in paper I–V. Structural analyses have been performed using a combination of nuclear magnetic resonance spectroscopy and chemical component analysis.

The elucidated structures in paper I–III, as well as paper V, are composed of linear repeating units of varying composition and length. In paper IV, the structure of the O-polysaccharide repeating unit of E. coli TD2158 is determined to be a branched hexasaccharide structure with a heterogeneous substitution pattern, with either a β-GlcpNAc or β-Glcp residue branching to the backbone chain. Incubation with bacteriophage HK620 tailspike protein shows that the polysaccharide is selectively cleaved at the α-GlcpNAc-(1→2)-α-Rhap-linkage of the backbone chain, yielding a 9:1 ratio of β-GlcpNAc/β-Glcp containing hexasaccharides after digestion.

In paper VI the conformational properties of a trisaccharide, which constitutes an internal epitope of the LeaLex hexasaccharide over-expressed on the surface of squamous lung cancer cells, have been analyzed using NMR spectroscopy and molecular dynamics simulations. The β-(1→3)-linkage of the trisaccharide was shown to be highly flexible.

Place, publisher, year, edition, pages
Stockholm: Department of Organic Chemistry, Stockholm University, 2012. 54 p.
NMR spectroscopy, structural analysis, conformational analysis, carbohydrates, polysaccharides
National Category
Organic Chemistry
Research subject
Organic Chemistry
urn:nbn:se:su:diva-75050 (URN)978-91-7447-457-2 (ISBN)
Public defence
2012-05-04, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: Submitted. Paper 6: Submitted.

Available from: 2013-04-09 Created: 2012-04-03 Last updated: 2013-11-18Bibliographically approved

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