Fusion proteins consisting of Bet v 1 and Phl p 5 form IgE-reactive aggregates with reduced allergenic activity
N. Najafi, G. Hofer, P. Gattinger, D. Smiljkovic, K. Blatt, A. Stöcklinger, W. Keller, P. Valent, J. Thalhamer, R. Valenta, S. Flicker., Scientific Reports, 9(1):4006. doi: 10.1038/s41598-019-39798-8.
Immunoglobulin E (IgE)-mediated allergy is the most common immunological hypersensitivity disease. The prevalence has been continuously rising over the last decades and currently, more than 25% of the population is affected. IgE-allergen complexes induce mast cell and basophil activation and thus immediate allergic inflammation.
Major birch and grass pollen allergens, Bet v 1 and Phl p 5 represent two of the most common allergens that induce strong allergic reactions in allergic patients as already repeatedly demonstrated by skin and nasal provocation testing.
To understand the potency of allergens to induce such severe allergic symptoms Dr. Nazanin Najafi from the group of Assoc. Prof. Sabine Flicker started to investigate different properties of the highly potent allergens, Phl p 5 and Bet v 1. In her study, which was recently published in Scientific Reports, a hybrid protein consisting of the complete mature sequence of Phl p 5 fused to the complete sequence of Bet v 1 was constructed, expressed and purified. Since each of these allergens occurs as soluble and monomeric protein, they expected the hybrid protein to remain fully IgE-reactive, allergenic and monomeric. Size exclusion experiments revealed that the majority of hybrid protein appears as high molecular weight aggregates of more than 1000 KDa. These results were in correspondence with illustrations obtained from negative stain electron microscopy. Hybrid, as an aggregated protein compared with equimolar mix of Phl p 5 and Bet v 1, exhibited a stronger IgE reactivity with sera from patients who were allergic to birch and grass pollen. However, to their surprise, basophil activation tests with above mentioned patients, demonstrated a reduced allergenic activity of hybrid compared with that of equimolar mix of the isolated allergens.
This study demonstrates clearly that a considerable reduction of allergenic activity can also be achieved through molecular aggregation of allergens not only by various recombinant DNA technologies such as mutation, reassembly, or fragmentation. Dr. Najafi explains the reduction of allergenic activity of the hybrid through a reorientation of the spatial arrangement of IgE epitopes in an unfavorable position for cross-linking of effector cell-bound IgE antibodies. Therefore the hybrid protein could be of clinical impact as controlled molecular aggregation may be used as a new technology for reducing the allergenic activity of allergens and thus for the engineering of a new type of allergy vaccines.
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