Identification of a Soybean Seed Biotinylated Protein as a Novel Allergen

RATIONALE: The aim of the study was to identify novel soy allergens. Although soybean allergens have been studied previously, a systematic search for additional allergenic proteins is required for the development of effective diagnostic and immunotherapeutic strategies.METHODS: A cDNA library from developing soybeans was constructed and screened for the presence of allergenic orthologs characterized in other seeds. The proteins encoded by these cDNAs were produced in a bacterial system and purified using affinity chromatography. IgE binding to the recombinant proteins was investigated by Western blotting and ELISA using 25 human sera containing IgEs to soy. Crude soybean and peanut protein extracts were also used in binding-inhibition assays to test the specificity of IgE binding to the candidate allergens.RESULTS: Our approach identified several candidate allergens, one of which referred to a seed specific biotinylated protein (SBP). Expression of a full length SBP cDNA produced a protein of approximately 70 kDa that was boiling resistant. More than 50% of the sera tested reacted with this rSBP in either immunoblots or ELISA. Inhibition of binding using crude soy extracts was greater than that observed when inhibition was carried out using peanut extracts. However, positive inhibition by peanut extracts suggests cross-reactivity between SBP from peanut and soy.CONCLUSIONS: The seed specific biotinylated from soy has been characterized as a novel allergen. Based on the percentage of sera recognized (>50%), this protein may represent a major allergen. The results from the inhibition experiments suggest that the peanut ortholog may also be allergenic. RATIONALE: The aim of the study was to identify novel soy allergens. Although soybean allergens have been studied previously, a systematic search for additional allergenic proteins is required for the development of effective diagnostic and immunotherapeutic strategies. METHODS: A cDNA library from developing soybeans was constructed and screened for the presence of allergenic orthologs characterized in other seeds. The proteins encoded by these cDNAs were produced in a bacterial system and purified using affinity chromatography. IgE binding to the recombinant proteins was investigated by Western blotting and ELISA using 25 human sera containing IgEs to soy. Crude soybean and peanut protein extracts were also used in binding-inhibition assays to test the specificity of IgE binding to the candidate allergens. RESULTS: Our approach identified several candidate allergens, one of which referred to a seed specific biotinylated protein (SBP). Expression of a full length SBP cDNA produced a protein of approximately 70 kDa that was boiling resistant. More than 50% of the sera tested reacted with this rSBP in either immunoblots or ELISA. Inhibition of binding using crude soy extracts was greater than that observed when inhibition was carried out using peanut extracts. However, positive inhibition by peanut extracts suggests cross-reactivity between SBP from peanut and soy. CONCLUSIONS: The seed specific biotinylated from soy has been characterized as a novel allergen. Based on the percentage of sera recognized (>50%), this protein may represent a major allergen. The results from the inhibition experiments suggest that the peanut ortholog may also be allergenic.