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An alternate pattern for globoside oligosaccharide expression in Haemophilus influenzae lipopolysaccharide: Structural diversity in nontypeable strain 1124
Södertörn University, School of Life Sciences. Karolinska Institutet.
National Research Council of Canada, Ottawa, Canada.
National Research Council of Canada, Ottawa, Canada.
John Radcliffe Hospital, Oxford, UK.
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2005 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 44, no 13, 5207-5224 p.Article in journal (Refereed) Published
Abstract [en]

Common structural motifs of Haemophilus influenzae lipopolysaccharide (LPS) are globotetraose [beta-D-GalpNAc-(1 -> 3)-alpha-D-Galp-(1 -> 4)-beta-D-Galp-(1 -> 4)-beta-D-Glcp] and its truncated versions globoside [alpha-D-Galp-(1 -> 4)-beta-D-Galp-(1 -> 4)-beta-D-Glcp] and lactose [beta-D-Galp-(1 -> 4)-beta-D-Glcp] linked to the tenninal heptose (HepIII) of the triheptosyl inner-core moiety L-alpha-D-Hepp-(1 -> 2)-[PEA -> 6]-L-alpha-D-Hepp-(1 -> 3)L-alpha-D-Hepp-(1 -> 5)-[PPEA -> 4]-alpha-Kdo-(2 -> 6)-lipid A. We report here structural studies of LPS from nontypeable H. influenzae strain 1124 expressing these motifs linked to both the proximal heptose (HepI) and HepIII at the same time. This novel finding was obtained by structural studies of LPS using NMR techniques and electrospray ionization mass spectrometry (ESI-MS) on O-deacylated LPS and core oligosaccharide material (OS) as well as ESI-MSn on permethylated dephosphorylated OS. The use of defined mutants allowed us to confirm structures unambiguously and understand better the biosynthesis of each of the globotetraose units. We found that lgtC is involved in the expression of beta-D-Galp-(1 -> 4)-beta-D-Galp in both extensions, whereas lic2A directs only the expression Of beta-D-Ga1p-(1 -> 4)-beta-D-Glcp when linked to HepIII. The LPS of NTHi strain 1124 contained sialylated glycoforms that were identified by CE-ESI-MS/MS. A common sialylated structure in H. influenzae LPS is sialyllactose linked to HepIII. This structure exists in strain 1124. However, results for the lpsA mutant indicate that sialyllactose extends from HepI as well, a molecular environment for sialyllactose in H. influenzae that has not been reported previously. In addition, the LPS was found to carry phosphoryleholine, O-linked glycine, and a third PEA group which was linked to O3 of HepIII.

Place, publisher, year, edition, pages
2005. Vol. 44, no 13, 5207-5224 p.
National Category
Biological Sciences
Identifiers
URN: urn:nbn:se:sh:diva-31883DOI: 10.1021/bi047480hISI: 000228099000025PubMedID: 15794658OAI: oai:DiVA.org:sh-31883DiVA: diva2:1069219
Available from: 2017-01-27 Created: 2017-01-27 Last updated: 2017-01-27Bibliographically approved
In thesis
1. Structural diversity of the lipid A and core oligosaccharide moieties of the lipopolysaccharides from nontypeable and serotype f Haemophilus influenzae
Open this publication in new window or tab >>Structural diversity of the lipid A and core oligosaccharide moieties of the lipopolysaccharides from nontypeable and serotype f Haemophilus influenzae
2005 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes structural studies of the oligosaccharide and lipid A moieties of lipopolysaccharides (LPSs) isolated from disease-causing Haemophilus influenzae strains. The nontypeable strains were clinical isolates from the middle ear of children suffering from otitis media and the serotype f strains had been collected from three adults with respiratory tract infections. The LPS molecules are situated on the cell wall of H. influenzae strains and they play a very important role in colonization, infection, evasion of host immune system and inflammatory response. Previous studies have implicated the heterogeneous repertoire of LPS structures within a strain and mimicry of human cell wall structures to be involved in the diseasecausing behavior of this organism. Structural analysis of the oligosaccharide moieties with advanced applications of nuclear magnetic resonance (NMR) and various electrospray ionization mass spectrometry (ESI-MS) techniques revealed novel structural features in each of the investigated strains. All of the strains displayed a very complex mixture of LPS structures that differed between and within the pathogens. Moreover, all of the strains had the capacity to express mimics of human glycolipids. The genetic basis for LPS biosynthesis for H. influenzae is established for the strain of which the complete genome has been determined. In this thesis the function of the genes involved in the biosynthesis of LPS was investigated in a nontypeable strain by using the combination of genetic engineering and structural analysis. The synergy of genomics and analytical carbohydrate chemistry led to the identification of novel structural epitopes, and furthermore, enabled us to identify a new function for one of these genes. The most recent structural study of lipid A from H. influenzae was conducted in 1988 on a mutant strain. The results of that study established the presence of only one lipid A structure. in this thesis we investigated lipid A from both nontypeable and serotype wild type strains by performing tandem ESI-MS and the results confirmed earlier findings but also evidenced other lipid A structures previously not associated with H. influenzae. Moreover, all of the strains exhibited a heterogeneous population of lipid A molecules.

Place, publisher, year, edition, pages
Stockholm: Karolinska instiutet, 2005. 60 p.
National Category
Biological Sciences
Identifiers
urn:nbn:se:sh:diva-31887 (URN)91-7140-504-6 (ISBN)
Public defence
2005-11-25, 4U Solen, Alfred Nobels allé 8, Huddinge, 09:00 (English)
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Available from: 2017-01-27 Created: 2017-01-27 Last updated: 2017-01-27Bibliographically approved

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