Molecular characterization of a cross-reactive Juniperus oxycedrus pollen allergen, Jun o 2: a novel calcium-binding allergen

J Allergy Clin Immunol. 1998 Jun;101(6 Pt 1):772-7. doi: 10.1016/S0091-6749(98)70306-9.

Abstract

Background: Species belonging to the Cupressaceae family are a relevant source of allergens that are present in a wide number of countries.

Objective: We sought to identify, purify, and characterize recombinant allergens from Juniperus oxycedrus, a species belonging to the Cupressaceae family.

Methods: Double-stranded cDNA was synthesized from mRNA and cloned into the lambda-ZAP expression vector. IgE screening of the library was performed with a pool of sera from subjects allergic to Cupressaceae. A recombinant 6xHis-tagged Juniperus oxycedrus allergen, Jun o 2, was expressed in Escherichia coli and purified by Ni2+ affinity chromatography. It was studied further by immunoblotting inhibition with pollen extracts from other Cupressaceae, Oleaceae, Urticaceae, and Graminaceae. The role of protein-bound calcium on the allergen's IgE-binding capacity was tested in a plaque assay in the presence or absence of EGTA.

Results: A cDNA coding for a newly identified Juniperus oxycedrus pollen allergen, rJun o 2, was isolated. The deduced amino acid sequence contained four typical Ca2+ binding sites and showed a significant sequence similarity to calmodulins. Depletion of Ca2+ in the plaque assay led to a loss of IgE-binding capacity of rJun o 2. Immunoblotting inhibition revealed that J. oxycedrus, J. ashei, Cupressus arizonica, C. sempervirens, Parietaria judaica, Olea europaea, and Lolium perenne pollen extracts were able to inhibit IgE binding to blotted rJun o 2 at different concentrations.

Conclusion: rJun o 2 contains IgE-binding epitopes shared by taxonomically unrelated species, and therefore it can be regarded as a new panallergen. These findings could contribute to an explanation for the phenomenon of multiple positive test results in polysensitized patients and the potential symptom-eliciting role of allergenic sources previously not encountered.

MeSH terms

  • Allergens / genetics*
  • Allergens / immunology*
  • Amino Acid Sequence
  • Antigens, Plant
  • Calcium-Binding Proteins / genetics*
  • Calcium-Binding Proteins / immunology*
  • Cloning, Molecular
  • Cross Reactions
  • DNA, Complementary / genetics
  • DNA, Complementary / isolation & purification
  • Escherichia coli
  • Gene Library
  • Humans
  • Juniperus
  • Molecular Sequence Data
  • Pollen*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Sequence Alignment

Substances

  • Allergens
  • Antigens, Plant
  • Calcium-Binding Proteins
  • DNA, Complementary
  • Jun o 2 allergen
  • Recombinant Proteins

Associated data

  • GENBANK/AF031471