ANTIGEN 5 FROM POLYBIA SCUTELLARIS, A NATURALLY OCURRING HYPOALLERGENIC VARIANT: EXPRESSION, CHARACTERIZATION AND CROSS-REACTIVITY WITH AN ALLERGENIC HOMOLOG.

S VINZÓN, C NOWICKI, E RIVERA, MJ BISCOGLIO

BUENOS AIRES, ARGENTINA

Congreso; XXI WORLD ALLERGY CONGRESS; 2009

WORLD ALLERGY ORGANIZATION

  Background: Antigen 5 (Ag5) is the main allergenic protein found in vespid venoms. However, results attained so far in our laboratory have shown that Ag5 from Polybia scutellaris (Poly s5), a South American wasp, is a hypoallergenic variant. The principal aim of this study is to further characterize this molecule in order to asses its usefulness as a tool in immunotherapy. As only low yields of the native Poly s5 (nPoly s5) can be obtained, it became relevant to express it as a recombinant protein.   Methods: nPoly s5, which is not glycosylated, was purified from the venom by HPLC. For heterologous expression and in order to preclude protein glycosylation, we performed a site-directed mutagenesis of 144Asn to Ser. The recombinant protein (mut rPoly s5) was then expressed in Pichia pastoris and purified to homogeneity. Proteins were injected to BALB/c mice and cross-reactivity with Ag5 from Polistes annularis (Pol a5), a well-known allergen, was determined by means of different immunological techniques.   Results: We found that mut rPoly s5 was successfully expressed in a non-glycosylated form. Inhibition IgG ELISA with the native variant showed that the recombinant protein maintains the immunological reactivity of the native protein. A significant cross-reactivity between Poly s5 and Pol a5 was detected at the IgG level by indirect ELISA using either the autologous or the homologous Ag5 as the coating agent and also as the booster protein in a non-standard immunization protocol. Nevertheless, we were able to find two differences between Poly s5 and Pol a5: on the one hand, they show a different degree of immunogenicity and, on the other hand, each has at least one different immunodominant epitope.   Conclusion: Our results show that both native and recombinant forms of Poly s5 are suitable hypoallergenic candidates for immunotherapy.