Molecular Differences in Major and Minor Allergens From US Grass Species Support the Association of Several Pollens for Specific Immunotherapy

Despite a high IgE cross-reactivity, pollen allergens display molecular differences between grass species, and consequently exhibit complementary T and B cell epitope repertoires. The extent of this molecular complementarity across distinct Pooideae species was evaluated for both major and minor pollen allergens. Group 1 and 5 pollen allergens from five grasses commonly distributed in the US, namely Orchard, Timothy, Ryegrass, Sweet Vernalgrass and Kentucky Bluegrass, were purified and characterized in terms of primary structure (using mass spectrometry, or MS), T cell reactivity (based on the induction of CD154 in PBMCs) and IgE binding affinities (assessed in ELISA-inhibition). Animal antibodies were raised against various allergens. Allergen-specific glycoepitopes were analyzed by 1D- and 2D-electrophoresis and Western blotting. Purified group 1 and 5 allergens exhibited considerable intra- and inter-species variability both in terms of aminoacid sequences and post-translational modifications. For most grass pollen-sensitized patients, this variability translates into T and/or B cell responses restricted to allergens from some of, but not all, species tested. Antibodies raised against group 2-4, 6, 11 or 12 allergens often react with species-restricted or -semi-restricted epitopes. Two-dimensional protein fractionation followed by MS confirmed that minor allergens display species-specific features. Species-specific glycoepitopes have been evidenced, and shown to elicit IgE in patients. Substantial differences are found between pollen allergens from five common US grasses, translating into species-restricted or -semi-restricted T and/or B cell epitopes. Combining those pollens for allergen-specific immunotherapy extends the repertoire of epitopes provided, and consequently should be more efficacious.