An allergenic polypeptide representing a variable region of hsp 70 cloned from a cDNA library of Cladosporium herbarum.

BACKGROUND

Extracts of Cladosporium herbarum, a major source of fungal aeroallergens, exhibit a complex profile of IgE-binding proteins. Yields of conventionally purified allergens from this mold have been insufficient to permit further molecular analyses.

OBJECTIVE

To enhance and simplify the purification of allergens from C. herbarum, we have sought to use recombinant DNA techniques to clone, identify and bacterially express immunoselected C. herbarum allergens.

METHODS

We constructed a cDNA library in lambda ZAP II using mRNA isolated from C. herbarum. From this library, phage clones encoding a new allergen were immunoselected using pooled human atopic IgE. The cloned cDNA was excised from the phage vector as a recombinant pBluescript II SK-phagemid and sequenced. Expression of the recombinant allergen was carried out in E. coli XL1-blue transformants of the phagemid. Bacterial lysates from cells induced to express the cloned allergen were immunoblotted and probed with individual human atopic IgEs.

RESULTS

The cDNA clone encodes a 278 amino acid polypeptide homologous to the C-terminal portion of 70 kDa heat shock protein (hsp 70). The polypeptide possesses features common to other hsps 70, i.e. a similar hydropathic profile and a variable C-terminal region with conserved sequence at the very C-terminus. Binding of the recombinant peptide to IgE from 38% of atopic sera or plasma from individuals allergic to C. herbarum was demonstrated.

CONCLUSION

These results indicate that amino acid substitutions are relatively conserved even in the variable C-terminal regions of hsp 70 species. Thus, this study should draw attention to the possibility of induction of anaphylactic responses in a sensitized individual when hsp 70 from any pathogenic species is administered for vaccination.