44G-10 |
Development of non-allergenic shrimp: Mutational analysis of the major B cell epitopes of the shrimp allergen tropomyosin (Pen a 1). |
R. AYUSO, G. Reese, and S. B. Lehrer. Section of Allergy and Immunology, Tulane University Health Sciences Center, 1700 Perdido St, 3 Floor, New Orleans, LA 70112 Of the food hypersensitivity reactions that affect approximately 8% of children and 2% of adults, shellfish (particularly crustaceans) cause a significant percentage of reactions. Since immunotherapy with food allergens can have severe side effects, and avoidance of the offending food is not always possible, a feasible approach would be to develop allergens with B cell epitopes modified to reduce their IgE binding capacity. The muscle protein tropomyosin is the only major shrimp allergen (Pen a 1); 8 IgE binding epitopes have been recently identified. The objective was to study the effect of amino acid substitution on reduction of the IgE binding capacity of those B cell epitopes was assessed in this study; IgE antibody reactivities of shrimp allergic subjects to the identified epitopes were compared with reactivities to mutated sequences substituted with amino acids from homologous sequences of non-allergenic vertebrate tropomyosins. All possible substitution combinations (up to 5 per peptide) based on the differences between Pen a 1 epitopes and homologous vertebrate sequences were tested using synthetic overlapping peptides (Genosys). Single amino acid substitutions may abolish, reduce, not alter or even increase IgE binding to the modified peptide, while two or more substitutions in general abolish IgE binding to the modified peptides. Non-conservative amino acid substitutions which are in the center of the epitope are more likely to abolish IgE binding than those which are conservative or in peripheral parts of the peptide. For example, substitution of Serine for Phenylalanine in position 269 of epitope 5b:(266-273), abolishes IgE binding to the mutated peptide in 4/4 subjects tested. Similar single amino acid substitutions that abolish IgE binding to the modified Pen a 1 peptides were identified in the other epitopes. These studies indicate that single amino acid substitutions can significantly reduce or abolish IgE antibody binding; this may be useful in future development of safer allergen vaccines or hypoallergenic foods.
Session 44G, Toxicology & Safety Evaluation
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