title: Studies on the carbohydrate moiety of Pla l 1 allergen. identification of a major N-glycan and significance for the immunoglobulin E-binding activity.
abstract: BACKGROUND
  Pla l 1, the major allergen of Plantago lanceolata pollen, is a glycoprotein that contains an N-glycosylation site. Carbohydrate moieties of many allergenic glycoproteins have been reported to be IgE-binding determinants responsible for cross-reactivity among different species.
  
  
  OBJECTIVE
  To identify the kind of linkages and the type of glycans present in Pla l 1 and to investigate their contribution to the allergic response to this allergen.
  
  
  METHODS
  Pla l 1 was deglycosylated by N-glycosidase A and the IgE-binding ability of the unglycosylated protein was evaluated by dot-blot. Identification of beta1 --> 2 xylose and/or alpha1 --> 3 fucose residues in Pla l 1 N-glycan was carried out by incubation with specific antibodies from rabbit antiserum against HRP (anti-HRP). The contribution of this N-glycan to total IgE reactivity was analysed quantitatively by pre-incubation of Pla l 1 with anti-HRP prior to incubation with sera. The role of the carbohydrate moiety of Pla l 1 in cross-reactivity was studied by RAST using unrelated glycoproteins with known sugar composition and structure.
  
  
  RESULTS
  The effectiveness of N-glycosidase A to deglycosylate Pla l 1 and the ineffectiveness of the treatment with PNGase F indicate that Pla l 1 carries a complex N-glycan with an alpha1 --> 3 fucose residue in its structure. Furthermore, the presence of beta1 --> 2 xylose and/or alpha1 --> 3 fucose residues was identified in this N-glycan by means of an ELISA. Pre-incubation of Pla l 1 with an anti-HRP antibody caused a weak but significant reduction in IgE reactivity. Some sera from P. lanceolata-allergic patients reacted positively with four glycoproteins that bear N-glycans of complex type but not with fetuine.
  
  
  CONCLUSIONS
  Pla l 1 is a glycoprotein that carries at least a complex, major N-linked glycan, with a alpha1 --> 3 fucose residue in its structure and probably also a beta1 --> 2 xylose. This glycan moiety does not seem to constitute a relevant allergenic epitope of Pla l 1.
authors.affiliation: Research and Development Department, ALK-Abelló, Madrid, Spain. belen.calabozo@es.alk-abello.com
authors.forename: B
authors.initials: B
authors.lastname: Calabozo
authors.forename: D
authors.initials: D
authors.lastname: Barber
authors.forename: F
authors.initials: F
authors.lastname: Polo
mesh_terms: D000485:Allergens / Q000737:chemistry* / Q000276:immunology; D000937:Antigen-Antibody Reactions; D002240:Carbohydrate Sequence; D003429:Cross Reactions; D006023:Glycoproteins / Q000737:chemistry; D007073:Immunoglobulin E / Q000276:immunology*; D010940:Plant Proteins; D010943:Plantago / Q000276:immunology*
chemical_list: D000485:Allergens; D006023:Glycoproteins; C473663:Pla l 1 allergen, Plantago lanceolata; D010940:Plant Proteins; D007073:Immunoglobulin E