Purification and characterization of a major cross-reactive allergen from Epicoccum purpurascens

Abstract

Background: Epicoccum purpurascens (formerly nigrum) (EP), is a ubiquitous saprophytic mould found both indoors and outdoors. Several studies have reported sensitization to EP in 5-7% of different populations worldwide. The diagnosis of mould allergy requires a standardized fungal extract that contains all its important allergenic proteins. The crude allergen extract from EP was standardized earlier, however none of its allergens have been purified.

Methods: A major allergen from spore-mycelia extract of EP was purified using concanavalin A (Con A) Sepharose chromatography, gel filtration and electro-elution. The allergen isolated was characterized for its IgE-binding ability and cross-reactivity with five well-known allergenic fungi by ELISA and immunoblot.

Results: A 33.5-kD glycoprotein allergen of EP, Epi p 1, was purified to homogeneity. All the EP allergic patients' sera tested recognized this protein. Periodate modification of Epi p 1 showed partial loss in IgE binding while proteinase K treatment caused complete loss in binding to IgE. Dose-dependent inhibition in binding of rabbit anti Epi p 1 antibodies was obtained with Epi p 1, Aspergillus fumigatus, Alternaria alternata, Curvularia lunata, Cladosporium herbarum and Fusarium solani in ELISA. Rabbit antibodies to all the above five fungi recognized Epi p 1 in immunoblot, confirming that Epi p 1 shares common epitopes with the fungi tested.

Conclusion: A major glycoprotein allergen of 33.5 kD was purified from EP which cross-reacts with other fungi. Hence this glycoprotein can be exploited to reduce the panel of allergen extracts used for therapy of mould allergy.