Molecular Characterization of Timothy Grass Pollen Group V Allergens

Abstract

Phl p V is the dominant allergen of timothy grass (Phleum pratense) with two isoforms having the apparent molecular weights of 38 (Phl p Va) and 32 kD (Phlp Vb) under Western blot conditions. Two-dimensional electrophoresis/immunoblotting reveals that each isoform is split into at least four isoallergens. Structural differences in the isoforms are shown by N-terminal sequencing (only 60% identity), by reaction patterns of monoclonal antibodies and, more convincingly, by enzymic degradation of purified isoforms followed by immunologic fingerprinting. These findings are confirmed by the deduced primary protein structure of cloned Phl p Va and Phlp Vb. Experiments with IgE – affinity-purified by immobilized recombinant allergens or their fragments – reveal identical epitopes and at least one different epitope between the isoforms. Furthermore, on Phlp Va we can localize different IgE-reactive epitopes at the C terminus as well as the N terminus. By probing serum from 11 patients on recombinant C- or N-terminal fragments, an individual reaction pattern was found. Testing the histamine liberation potency of the fragments, we found the N-terminal fragment of Phlp Va to be superior to that of the C-terminal fragment or the whole molecule. These results give insights into the variability of allergens, the individuality of human reaction patterns to epitopes and the alteration of allergenicity to higher or lower levels by fragmentation.

Identification and Characterization of the Allergenic Proteins of Bahia Grass <i>(Paspalum notatum)</i> Pollen

“…Like many other grass allergens, pollen of Bahia grass contains many different proteins but only a few are allergenic proteins [7, 8, 16, 17, 18, 19]. These allergenic proteins of Bahia grass have molecular weights of 28, 31, 33, 45 kD and similar isoelectric points within the pI range of 6.39–7.05.…”

“…Because all 4 allergens were present in similar proportions in the same IEF fractions and focused in the same pI range, they may represent 1 or 2 large proteins subdivided into 4 isoallergens. A major allergen of timothy grass pollen, Ph1 p 5, occurs as 2 isoforms that split into at least 4 isoallergens [16]. Alternatively, the 4 polypeptides of Bahia or timothy grass may represent 4 different isoforms with same pI values.…”

C-Terminal Cysteine Residues Determine the IgE Binding ofAspergillus fumigatusAllergen Asp f 2

“…The mediators released as a result of allergen-induced crosslinking of IgE Abs on mast cell and basophil receptors are mainly responsible for the clinical symptoms of type 1 allergy. The structural analysis of allergens is of utmost importance because only a small number of Ags are allergens and the characteristic structural features of these proteins are essential in giving rise to symptoms in allergic disorders (1). Among various inhalant allergens, fungal spores are universally recovered both indoors and outdoors and are recognized as an important cause of respiratory allergy (2)(3)(4)(5).…”

From the allergen-recognition by antibodies to new therapeutic concepts

“…Diese beiden Pollenproteine stellen daher wesentliche allergene Komponenten dar, und sie sind wichtige Ziele für die Immuntherapie. Mit Hilfe der Mimotop-Technologie konnten wir kürzlich die IgE-Epitope von Bet v 1 und einer wichtigen Isoform von Phl p 5, dem Phl p 5a [35], identifizieren [22,36]. Wir verwendeten affinitätsgerei-nigtes Patienten-IgE, um damit Liganden aus einer zirkulären dekameren Peptidbibliothek [29] zu selektionieren.…”