Project Number: 6435-43440-044-00
Project Type: Appropriated

Start Date: Oct 01, 2010
End Date: Sep 30, 2015

Objective:
Objective 1: Identify and compare the structural, chemical, functional, and immunological characteristics of peanut with homologous; less allergenic legume (green pea and soy) and tree nut allergens in raw and processed forms towards delineating the clinically-relevant antibody-allergen interactions. Objective 2: Clone, express, and purify the major peanut and select tree nut and legume allergens, and fragments thereof, in recombinant form to further delineate clinically-relevant antibody-allergen interactions. Objective 3: Assess the role of processing-induced chemical or structural modifications on the individual allergens by systematically altering amino acids thought to be important in clinically-symptomatic allergic reactions. Objective 4: Develop computer models and/or determine NMR/crystal structures of native and recombinant peanut and select tree nut and legume allergens in raw and processed forms. Objective 5: Combine the structural information obtained with the empirical knowledge from Objectives 1, 2, and 3 to identify clinically-relevant allergen-antibody interactions in peanut and tree nut allergy. Objective 6: Develop processing technologies for peanut products with reduced allergenic properties. Objective 7: Develop and improve immunoassays for detection of peanut, select tree nut, and soy allergen residues before and after processing (i.e. roasting, baking into cookies, etc.). Objective 8: After establishing standardized protocols for determining threshold doses for peanut, select tree nut, and soy allergens, determine threshold (minimal eliciting) dose of reactivity for processed forms of peanut allergens and develop computational and statistical models to estimate population thresholds.

Approach:
Specifically, peanuts and tree nuts will be subjected to thermal processing (i.e., roasting). New allergens or changes in allergenic properties of existing allergens due to the thermal processing will be identified by immunoassays, using serum (containing IgE antibodies) from peanut and/or tree nut allergic individuals. Proteins found to be immunologically altered by thermal processing will be purified by conventional chromatography and analyzed for alterations in size, structure, digestibility, and binding to various antibodies, including anti-Maillard reaction products and specific anti-allergen antibodies. The specific amino acid residues, thought to be modified during different processing events and to contribute to altered allergenic properties (i.e. IgE binding), will be identified. These amino acids will be identified by cloning and expression of select recombinant major allergens of peanut and tree nuts in E. coli followed by site directed mutagenesis, simulated processing and immunological analysis of previously identified IgE binding sites, specifically, sites thought to be modified by processing. Understanding the molecular basis of processing-induced alterations of allergens will guide development of processing technologies towards reduced allergenicity of nuts. This knowledge will also contribute to the development of better labeling practices and detection tools for industry and regulatory agencies resulting in better protection of consumers.