Molecular characterization and allergenic activity of Lyc e 2 (β-fructofuranosidase), a glycosylated allergen of tomato

Sandra Westphal, Daniel Kolarich, Kay Foetisch, Iris Lauer, Friedrich Altmann, Amedeo Conti, Jesus F. Crespo, Julia Rodríguez, Ernesto Enrique, Stefan Vieths, Stephan Scheurer*

*Corresponding author for this work

Research output: Contribution to journalArticle

90 Citations (Scopus)

Abstract

Until now, only a small amount of information is available about tomato allergens. In the present study, a glycosylated allergen of tomato (Lycopersicon esculentum), Lyc e 2, was purified from tomato extract by a two-step FPLC method. The cDNA of two different isoforms of the protein, Lyc e 2.01 and Lyc e 2.02, was cloned into the bacterial expression vector pET100D. The recombinant proteins were purified by electroelution and refolded. The IgE reactivity of both the recombinant and the natural proteins was investigated with sera of patients with adverse reactions to tomato. IgE-binding to natural Lyc e 2 was completely inhibited by the pineapple stem bromelain glycopeptide MUXF (Manα1-6(Xy1β1-2)Manβ1-4GlcNAcβ1-4(Fucα1-3) GlcNAc). Accordingly, the nonglycosylated recombinant protein isoforms did not bind IgE of tomato allergic patients. Hence, we concluded that the IgE reactivity of the natural protein mainly depends on the glycan structure. The amino acid sequences of both isoforms of the allergen contain four possible N-glycosylation sites. By application of MALDITOF mass spectrometry the predominant glycan structure of the natural allergen was identified as MMXF (Manα1-6 (Manα1-3)(Xy1β1-2)Manβ1-4GlcNAcβ1-4(Fucα1-3) GlcNAc). Natural Lyc e 2, but not the recombinant protein was able to trigger histamine release from passively sensitized basophils of patients with IgE to carbohydrate determinants, demonstrating that glycan structures can be important for the biological activity of allergens.

Original languageEnglish
Pages (from-to)1327-1337
Number of pages11
JournalEuropean Journal of Biochemistry
Volume270
Issue number6
DOIs
Publication statusPublished - Mar 2003

Keywords

  • Food allergen
  • Glycoprotein
  • IgE reactivity
  • Lyc e 2
  • Tomato

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