Methods: IgE-reacting F. proliferatumcomponents were identified by immunoblot. Characterization of allergens and determination of IgE cross-reactivity were performed by cDNA cloning and immunoblot inhibition studies.
Results: The 36.5 kDa IgE-binding component of F. proliferatum reacted with a monoclonal antibody FUM20 against fungal vacuolar serine protease allergens. The cDNA of F. proliferatum vacuolar serine protease (Fus p 9.0101) was cloned. Nine serum samples known previously IgE-binding against the vacuolar serine protease allergen of Penicillium chrysogenum (Pen ch 18) showed IgE immunoblot reactivity against rFus p 9.0101. The purified rFus p 9.0101 can inhibit IgE and FUM20-binding against the 36.5 kDa component of F. proliferatum. Thus, a novel and important F. proliferatum vacuolar serine protease allergen (Fus p 9.0101) was identified. The rPen ch 18 can inhibit IgE-binding against rFus p 9.0101. It indicates that IgE cross-reactivity between the Fusarium and the Penicilliumvacuolar serine protease allergens also exists. Both rFus p 9.0101 K88A and rPen ch 18 K89A mutants did not inhibit IgE-binding against rFus p 9.0101. Lys88 was considered a critical core amino acid in IgE-binding against Fus p 9.0101 and a residue responsible for IgE cross-reactivity between Fus p 9.0101 and Pen ch 18 allergens.
Conclusions: Our results indicate vacuolar serine protease is an important allergen of F. proliferatum and an IgE cross-reactive major pan-fungal allergen.