Evaluating the potential allergenicity of dietary proteins using model strong to non-allergenic proteins in germ-free mice.

Currently no validated animal model is predictive of human responses in ranking purified dietary proteins in the prevalence or potency of food allergy in humans. Since the gastrointestinal microbiota is thought to influence oral tolerance, we hypothesize that a germ-free mouse model will more accurately predict atopic human responses than conventional mice. Germ-free C3H/HeN mice were immunized with 60 µg Ara h 2, BLG, or LOX by three weekly intraperitoneal (IP) injections with alum adjuvant. One week following the final immunization an IP challenge of 500 µg of Ara h 2, BLG, or LOX was administered. Thirty minutes post-challenge clinical scores were graded and body temperatures recorded. The presence of protein-specific IgE and mast cell protease concentrations in mouse sera were determined using ELISA. Upon challenge germ-free mice sensitized with Ara h 2 and BLG exhibited significantly more severe clinical scores compared to germ-free mice immunized with LOX. Hypothermic responses in challenged mice differed between the three proteins post-challenge. Results indicate that this model can differentiate between potent and non-allergens based on temperature drop, clinical scores, and biomarkers. Additional proteins with known human exposure and allergenicity are needed to confirm the predictive accuracy.

Evaluating Potential Risks of Food Allergy and Toxicity of Soy Leghemoglobin Expressed in Pichia pastoris.

SCOPE: The Soybean (Glycine max) leghemoglobin c2 (LegHb) gene was introduced into Pichia pastoris yeast for sustainable production of a heme-carrying protein, for organoleptic use in plant-based meat. The potential allergenicity and toxicity of LegHb and 17 Pichia host-proteins each representing ≥1% of total protein in production batches are evaluated by literature review, bioinformatics sequence comparisons to known allergens or toxins, and in vitro pepsin digestion. METHODS AND RESULTS: Literature searches found no evidence of allergenicity or toxicity for these proteins. There are no significant sequence matches of LegHb to known allergens or toxins. Eleven Pichia proteins have modest identity matches to minor environmental allergens and 13 Pichia proteins have significant matches to proteins from toxic sources. Yet the matched allergens and toxins have similar matches to proteins from the commonly consumed yeast Saccharomyces cerevisiae, without evidence of food allergy or toxicity. The demonstrated history of safe use indicates additional tests for allergenicity and toxicity are not needed. The LegHb and Pichia sp. proteins were rapidly digested by pepsin at pH 2. CONCLUSION: These results demonstrate that foods containing recombinant soy LegHb produced in Pichia sp. are unlikely to present an unacceptable risk of allergenicity or toxicity to consumers.