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Used in many ways, almonds (Prunus dulcis) are valued for their nutritional and sensory properties and are globally popular. The US accounts for over 80% of the global almond production. Although safely enjoyed by many, sensitive individuals experience adverse reactions to food upon exposure to the offending food. Allergy is one such adverse reaction. <p>The "big 8" foods, responsible for over 90% of food allergies are milk, eggs, soybeans, shellfish, crustaceans fish, wheat, peanuts, and tree nuts. Type I hypersensitivity diseases, such as food allergies, allergic asthma, rhinitis, skin inflammation, and anaphylactic shock, are mediated by immunoglobulin E (IgE). Type I food allergies are typically caused by food proteins. Symptoms of food allergies may range from allergic rhinitis (sinus inflammation) or minor skin rash to life-threatening anaphylaxis (respiratory collapse and cardiovascular collapse). <p> In the US, most food-induced anaphylactic reactions are attributed to peanut, tree nut, and shell fish with annual fatalities of ~150-200. <p>Allergies to tree nuts and peanuts can be severe, are generally not outgrown. Almond allergy has been identified as the 3rd most frequent tree nut allergy in the US and severe anaphylaxis to almond has been reported. <p>The portion of allergenic protein molecule responsible for IgE binding is known as the epitope. When the allergenic epitope cross-links two IgE molecules on the surface of basophils/mast cells, several mediator molecules are released that lead to an allergic reaction. Two types of epitopes on an allergen are recognized. The linear stretches of amino acids-linear epitope and conformational motif- the conformational epitope. Linear, conformational, or both type of epitopes may be present on an allergen. <p>Any treatment that denatures allergenic epitopes would be expected to reduce or eliminate IgE-mediated cross-linking on the basophils/mast cell surfaces and thus reduce or eliminate allergenicity. Such treatments may lead to the development of hypoallergenic food products. <p>The main project goal is to assess suitability of processing methods to develop hypoallergenic almonds and almond products. We have targeted four allergenic almond proteins 2S, 7S, 11S, and profilin for the proposed investigation. <p>Specific Objectives: <ol> <LI> Characterize almond seed proteins for their molecular properties. <LI>Determine immunodominant epitopes in the key allergenic proteins: 2S albumin, vicilin (7S protein), amandin (11S protein), and profilin. <LI> Assess the stability of immunodominant epitopes of select proteins in native and processed almonds. <LI> Assess immunodominant epitope stability in almonds subjected to in vitro proteolysis under simulated digestion conditions. </ol>We anticipate identification of almond sensitive patient sera IgE reactive epitopes on the targeted proteins. With suitable processing methods, partial or complete destruction of some of the IgE-reactive epitopes is expected. Together, these results are expected to provide basis for developing methods to produce hypoallergenic almonds and almond products.
Non-Technical Summary: The proposed investigation aims to: 1. Identify key allergenic proteins in almond seeds. 2. Determine motifs- linear stretches as well as three dimensional structural portions of the key allergens identified in (1) that are responsible almond induced allergies in sensitive individuals. 3. Design suitable food processing methods to partially or completely inactivate the motifs identified in (2) to help develop hypoallergenic almonds and almond products. <P> Approach: The overall goal of the proposed research is two-fold: a) To identify immunodominant epitopes in select almond allergens and b) To develop processes to render almonds hypoallergenic. Based on the ongoing investigations in our laboratories and those reported by others, four allergenic proteins in almond seeds appear to be responsible for almond allergies in patients. Of the four, three proteins- a legumin, also known as amandin, 11S, or AMP, a vicilin also referred to as a 7S protein, and a 2S albumin protein, are major allergens. The fourth, a profilin is a minor allergen. Using the cDNA clones from almond cDNA library, we plan to express and sequence the selected clones encoding for 11S, 7S, 2S, and profilin. Based on the cDNA derived amino acid sequence of each protein, we will synthesize overlapping peptides, 15 amino acids per peptide with 3 amino acids offset in each, covering the full length sequence of the target protein and identify the linear epitopes. Using mouse monoclonal antibodies (mAbs), the recombinant and native proteins, patient sera IgE, and immunoblotting techniques, we will identify the IgE reactive epitopes. Proteins from almond seeds and almonds products, processed and unprocessed, will be probed with select mAbs directed against targeted epitopes and patient sera IgE to identify hypoallergenic products and to assess stability of immunodominant epitopes.