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


OBJECTIVE
IgE-reactive B-cell epitopes of C herbarum enolase were analyzed, and cross-reactivity between fungal enolases was investigated.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Simon-Nobbe
B
B
Institute of Genetics and General Biology, University of Salzburg, Salzburg, Austria.
Probst
G
G
Kajava
A V
AV
Oberkofler
H
H
Susani
M
M
Crameri
R
R
Ferreira
F
F
Ebner
C
C
Breitenbach
M
M
D000485:Allergens / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D000528:Alternaria / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D000908:Antibodies, Fungal / Q000276:immunology*; D000946:Antigens, Fungal / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D002963:Cladosporium / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D003001:Cloning, Molecular; D018604:Epitope Mapping / Q000379:methods; D018985:Epitopes, B-Lymphocyte / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology*; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D010751:Phosphopyruvate Hydratase / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D017422:Sequence Analysis, DNA / Q000379:methods
D000485:Allergens; D000908:Antibodies, Fungal; D000946:Antigens, Fungal; D018985:Epitopes, B-Lymphocyte; D010455:Peptides; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E; D010751:Phosphopyruvate Hydratase