NATIVE AND RECOMBINANT FEL-DI AS PROBES INTO THE RELATIONSHIP OF ALLERGEN STRUCTURE TO HUMAN IGE IMMUNOREACTIVITY

To delineate the relationship between the structural conformation and the stability of an allergen and its antigenicity, we have chosen the major allergen from cat dander, Fel dl. From protein sequence analysis data we have examined the structure of the naturally occurring Fel dI and we have found it to exist as an anti-parallel heterodimer. We hav e used ELISA, RAST, Western blot and histamine release techniques to c ompare the IgE reactivity of a set of cat allergic patient samples to purified, native Fel dI and the E. coli expressed chains 1 and 2. Resu lts from these studies demonstrate a significant level of IgE reactivi ty to all forms when examined for direct binding. However, both blot a nd ELISA competition assays show a much higher reactivity to Fel dI in solution compared to the separate recombinant chains and this is supp orted by the histamine release data. Although native Fel dI chain 2 co ntains an N-linked carbohydrate moiety, this does not seem to play a r ole in the reactivity of IgE to chain 2. Denaturation of Fel dI with a lkali conditions leads to a dramatic decrease in IgE reactivity, even though measurable changes to the backbone structure of the protein are minimal. One proposed explanation is that both chains possess a core region determined by their primary structures and that the major IgE e pitopes are dependent upon them. The relative reactivity amongst these allergen forms varied with the method of analysis, implying that the conformational requirements for IgE antibody binding are best studied by the application of more than one experimental protocol. Results fro m these qualitative analyses afford insight into the allergenicity of this exceptionally stable cat pelt protein.