IgE-binding epitopes of enolases, a class of highly conserved fungal allergens.

Abstract

Cladosporium herbarum and Alternaria alternata are two of the most prominent fungal species inducing type I allergy. Previously, we have demonstrated that enolase (Cla h 6) is the second most important allergen of C herbarum in terms of frequency of sensitization. IgE-reactive B-cell epitopes of C herbarum enolase were analyzed, and cross-reactivity between fungal enolases was investigated. Cla h 6 glutathione-S-transferase fusion peptides were constructed by means of PCR cloning. A alternata enolase (Alt a 5) was isolated by screening a complementary (c)DNA expression library with a C herbarum enolase DNA probe. Mapping of Cla h 6 IgE-binding epitopes identified a peptide with a length of 69 amino acids (peptide 9), which bound IgE from 8 of 8 patients. Analysis of the conformation of peptide 9 revealed that it does not form a compact structure but rather spans the whole length of the protein, with side chains exposed to solvent at 3 locations. Peptide 9 in the context of Escherichia coli glutathione-S-transferase not only binds IgE but also competitively inhibits IgE binding to Alt a 5. This result indicates that the epitope or epitopes on peptide 9 constitute a major cross-reacting epitope or epitopes on the enolases from C herbarum and A alternata in the case of the one patient tested. We demonstrated that the glycolytic enzyme enolase is an allergen not only in C herbarum but also in A alternata. Additionally, enolase was shown to exhibit high cross-reactivity to other fungal enolases. On the basis of the results presented here, we propose the use of recombinant Cla h 6 or maybe even peptide 9 of Cla h 6 for diagnosis and possibly therapy of mold allergy.