Protein quaternary structure and aggregation in relation to allergenicity

In order to induce systemic food allergic reactions in humans, proteins after digestion in the human gastro-intestinal tract should still be able to bind IgE. The aim of the work presented in this thesis was to determine the effects of heating on the structure and digestibility of cupin and prolamin food allergens from peanuts, Brazil nuts and soybeans and to determine the effects of digestion on the IgE binding capacity of these allergens. Representative allergen preparations were purified prior to the investigations. To this end, a large-scale purification method was developed for the purification of Brazil nut allergen Ber e 1, comprising its complete isoform pool. The denaturation temperature of the latter allergen at the common pH values of foods (5-7) appeared to be very high. As a result, this protein is assumed not to be denatured during common food processing and to be digested (mainly) in its native form. The native form of Ber e 1 is known to have a high stability towards peptic digestion, which likely explains the allergenicity of this protein. The representative quaternary structure of vicilin allergen Ara h 1 from peanuts appeared to be an oligomeric structure, in which the protein is assumed to interact with proanthocyanidins via a specific distribution of proline residues on its surface. Heat-induced aggregation prior to peptic hydrolysis under <i>in vitro</i> conditions did not affect the digestibility of Ara h 1, whereas heat-induced aggregation of legumin allergens Ara h 3 from peanuts and glycinin from soybeans (slightly) decreased the digestion rates. Regardless of pre-heating, the IgE binding capacity of the latter allergens diminished fast during pepsin digestion. As a result, in terms of systemic food allergic reactions, legumin allergens from peanuts and soybeans might not be very important allergens. Vicilin allergen Ara h 1 could still bind IgE after prolonged peptic digestion. Two pepsin-resistant epitopes were deduced and indications for four additional pepsin-resistant epitopes are presented. These epitopes are all situated on the unique N-terminal part of the allergen, which might explain why allergic reactions to peanuts are often severe, compared to other legume allergies