Gain of structure and IgE epitopes by eukaryotic expression of the major Timothy grass pollen allergen, Phl p 1

Tanja Ball, William Edstrom, Ludwig Mauch, Jacky Schmitt, Bernd Leistler, Helmut Fiebig, Wolfgang R. Sperr, Alexander W. Hauswirth, Peter Valent, Dietrich Kraft, Steven C. Almo, Rudolf Valenta

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Approximately 400 million allergic patients are sensitized against group 1 grass pollen allergens, a family of highly cross-reactive allergens present in all grass species. We report the eukaryotic expression of the group 1 allergen from Timothy grass, Phl p 1, in baculovirus-infected insect cells. Domain elucidation by limited proteolysis and mass spectrometry of the purified recombinant glycoprotein indicates that the C-terminal 40% of Phl p 1, a major IgE-reactive segment, represents a stable domain. This domain also exhibits a significant sequence identity of 43% with the family of immunoglobulin domain-like group 2/3 grass pollen allergens. Circular dichroism analysis demonstrates that insect cell-expressed rPhl p 1 is a folded species with significant secondary structure. This material is well behaved and is adequate for the growth of crystals that diffract to 2.9 Å resolution. The importance of conformational epitopes for IgE recognition of Phl p 1 is demonstrated by the superior IgE recognition of insect-cell expressed Phl p 1 compared to Escherichia coli-expressed Phl p 1. Moreover, insect cell-expressed Phl p 1 induces potent histamine release and leads to strong up-regulation of CD203c in basophils from grass pollen allergic patients. Deglycosylated Phl p 1 frequently exhibits higher IgE binding capacity than the recombinant glycoprotein suggesting that rather the intact protein structure than carbohydrate moieties themselves are important for IgE recognition of Phl p 1. This study emphasizes the important contribution of conformational epitopes for the IgE recognition of respiratory allergens and provides a paradigmatic tool for the structural analysis of the IgE allergen interaction.

Original languageEnglish (US)
Pages (from-to)217-227
Number of pages11
JournalFEBS Journal
Volume272
Issue number1
DOIs
StatePublished - Jan 2005

Fingerprint

Phleum
Pollen
Allergens
Immunoglobulin E
Epitopes
Poaceae
Insects
Glycoproteins
Proteolysis
Basophils
Baculoviridae
Histamine Release
Circular Dichroism
Crystallization
Structural analysis
Escherichia coli
Histamine
Mass spectrometry
Immunoglobulins
Mass Spectrometry

Keywords

  • Allergen
  • Allergy
  • Epitope
  • Eukaryotic expression
  • Phl p 1

ASJC Scopus subject areas

  • Biochemistry

Cite this

Ball, T., Edstrom, W., Mauch, L., Schmitt, J., Leistler, B., Fiebig, H., ... Valenta, R. (2005). Gain of structure and IgE epitopes by eukaryotic expression of the major Timothy grass pollen allergen, Phl p 1. FEBS Journal, 272(1), 217-227. https://doi.org/10.1111/j.1432-1033.2004.04403.x

Gain of structure and IgE epitopes by eukaryotic expression of the major Timothy grass pollen allergen, Phl p 1. / Ball, Tanja; Edstrom, William; Mauch, Ludwig; Schmitt, Jacky; Leistler, Bernd; Fiebig, Helmut; Sperr, Wolfgang R.; Hauswirth, Alexander W.; Valent, Peter; Kraft, Dietrich; Almo, Steven C.; Valenta, Rudolf.

In: FEBS Journal, Vol. 272, No. 1, 01.2005, p. 217-227.

Research output: Contribution to journalArticle

Ball, T, Edstrom, W, Mauch, L, Schmitt, J, Leistler, B, Fiebig, H, Sperr, WR, Hauswirth, AW, Valent, P, Kraft, D, Almo, SC & Valenta, R 2005, 'Gain of structure and IgE epitopes by eukaryotic expression of the major Timothy grass pollen allergen, Phl p 1', FEBS Journal, vol. 272, no. 1, pp. 217-227. https://doi.org/10.1111/j.1432-1033.2004.04403.x
Ball, Tanja ; Edstrom, William ; Mauch, Ludwig ; Schmitt, Jacky ; Leistler, Bernd ; Fiebig, Helmut ; Sperr, Wolfgang R. ; Hauswirth, Alexander W. ; Valent, Peter ; Kraft, Dietrich ; Almo, Steven C. ; Valenta, Rudolf. / Gain of structure and IgE epitopes by eukaryotic expression of the major Timothy grass pollen allergen, Phl p 1. In: FEBS Journal. 2005 ; Vol. 272, No. 1. pp. 217-227.
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