Helicobacter pylori cell surface interactions with glycosaminoglycans. Identification and characterisation of proteins binding to heparin/heparan sulphate.

Research output: ThesisDoctoral Thesis (compilation)

Abstract

Helicobacter pylori is a gastric pathogen which cause chronic type B gastritis and peptic ulcer disease. H. pylori produces virulence factors such as urease, vacuolating cytotoxin VacA, cag pathogenicity island-associated proteins, flagella and adhesins. These proteins interact with several host cell molecules such as sialylated cell surface glycoproteins, some of glycolipids and mucins. Glycosaminoglycans are widely distributed molecules in human cells and extracellular matrix and interact with H. pylori cell surface proteins. Heparan sulphate shows a pH-dependent binding to all strains, which is inhibited by sulphated glycosaminoglycans and other polysulphated molecules such as dextran sulphate and fucoidan. Using bioinformatics methods, the binding of H. pylori VacA cytotoxin to heparin and heparan sulphate was predicted. Specific peptides were synthesised and their interactions with surface-immobilised heparan sulphate studied in a BIAcore biosensor. Synthetic peptides bound to heparan sulphate with a moderate affinity. It was shown that native toxin interacts with immobilised heparin. Heparan sulphate was proposed as a receptor/co-receptor for VacA cytotoxin binding to host cell surfaces. H. pylori urease binds to surface-immobilised heparin and heparan sulphate. The interaction was characterised using a BIAcore biosensor. The binding was a thousand-fold higher at pH 5.5 compared to pH 6.5. The binding epitopes were identified by SELDI-TOF MS technology. Using a proteomic technology, four immunogenic heparin-binding proteins were identified: cell binding factor 2, urease, one outer membrane protein and one hypothetical protein. The binding of three of these proteins to heparin was demonstrated for the first time. The cell binding factor was identified as a specific and highly immunogenic H. pylori surface protein. 2-DE and 2-D immunoblotting allow efficient separation and identification of immunogenic proteins. Understanding how GAGs interact with pathogens, will allow the design of new experimental putative anti-microbial compounds based on sulphated polysaccharide structures.

Details

Authors
  • Meeme Utt
Organisations
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Microbiology in the medical area

Keywords

  • SELDI-TOF MS, bioinformatics, proteomics, immunoblotting, 2-DE, biosensor, heparan sulphate, heparin, binding, Helicobacter pylori, cell surface, bakteriologi, Mikrobiologi, mycology, bacteriology, Microbiology, mykologi, virologi, virology
Original languageEnglish
QualificationDoctor
Awarding Institution
Supervisors/Assistant supervisor
  • [unknown], [unknown], Supervisor, External person
Award date2002 Apr 3
Publisher
  • Meeme Utt, MMDI, University of Lund, Sölvegatan 23, S-223 62 Lund, Sweden,
Print ISBNs91-628-5158-6
Publication statusPublished - 2002
Publication categoryResearch

Bibliographic note

Defence details Date: 2002-04-03 Time: 13:15 Place: Rune Grubb-Salen, BMC, Lund External reviewer(s) Name: Magnusson, Karl-Eric Title: Professor Affiliation: Faculty of Health Sciences, Division of Medical Microbiology, University of Linköping, Sweden --- Article: I Meeme Utt and Torkel Wadström. Identification of heparan sulphate binding surface proteins of Helicobacter pylori: Inhibition of heparan sulphate binding with sulphated carbohydrate polymers. J. Med. Microbiol.1997, 46, 541-546. Article: II Meeme Utt, Bengt Danielsson and Torkel Wadström. Helicobacter pylori vacuolating cytotoxin binding to a putative cell surface receptor, heparan sulphate, studied by surface plasmon resonance.FEMS Immunology and Medical Microbiology, 2001, 30, 109-113. Article: III Meeme Utt, Bengt Danielsson, Jim McGuire and Torkel Wadström. Characterisation of Helicobacter pylori urease binding to heparin/heparan sulphate.Submitted. Article: IV Meeme Utt, Ingrid Nilsson, Åsa Ljungh and Torkel Wadström. Identification of novel immunogenic proteins of Helicobacter pylori by proteome technology.J. Immunol. Methods, 2002, 259, 1-10.