Three-Dimensional Structure of the Cross-Reactive Pollen Allergen Che a 3: Visualizing Cross-Reactivity on the Molecular Surfaces of Weed, Grass, and Tree Pollen Allergens

Abstract

Two EF-hand calcium-binding allergens (polcalcins) occur in the pollen of a wide variety of unrelated plants as highly cross-reactive allergenic molecules. We report the expression, purification, immunological characterization, and the 1.75-Å crystal structure of recombinant Che a 3 (rChe a 3), the polcalcin from the weed Chenopodium album. The three-dimensional structure of rChe a 3 resembles an α-helical fold that is essentially identical with that of the two EF-hand allergens from birch pollen, Bet v 4, and timothy grass pollen, Phl p 7. The extensive cross-reactivity between Che a 3 and Phl p 7 is demonstrated by competition experiments with IgE Abs from allergic patients as well as specific Ab probes. Amino acid residues that are conserved for the two EF-hand allergen family were identified in multiple sequence alignments of polcalcins from 15 different plants. Next, the three-dimensional structures of rChe a 3, rPhl p 7, and rBet v 4 were used to identify conserved amino acids with high surface exposition to visualize surface patches as potential targets for the polyclonal IgE Ab response of allergic patients. The essentially identical three-dimensional structures of rChe a 3, rPhl p 7, and rBet v 4 explain the extensive cross-reactivity of allergic patients IgE Abs with two EF-hand allergens from unrelated plants. In addition, analyzing the three-dimensional structures of cross-reactive Ags for conserved and surface exposed amino acids may be a first approach to mapping the conformational epitopes on disease-related Ags that are recognized by polyclonal patient Abs.

Allergenic Differences among Pollens of Three <i>Salsola</i> Species

“…Cross-reactivity between Chenopodiaceae pollens and other pollens, including grasses or trees, has been reported to be mediated by common allergens [18,19] or panallergens present in many different species [20]. In agreement with our clinical observations, an association between S. oppositifolia and O. europaea may be suggested.…”

Differences among Pollen-Allergic Patients with and without Plant Food Allergy

“…In this study we have analyzed 2 well-defined pollen-allergic patient groups and tested a wide and well-documented panel of purified allergens [10,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39]. Clinical data were compared with both the results obtained with the specific IgE responses to this panel and with a battery of commercially available pollen extracts used for skin tests.…”

“…The following natural or recombinant allergens, isolated by previously described methods, were included in the diagnostic panel: Phleum pratense: nPhl p 1 [19] and nPhl p 5 [20]; Cynodon dactilon: nCyn d 1 [21]; A. vulgaris: nArt v 1 [22]; O. europaea: nOle e 1 [23] and Ole e 9, as a mix of rN-terminal domain and rC-terminal domain from Ole e 9 [24,25]; P. lanceolata: nPla l 1 [26]; P. judaica: nPar j 1 [27]; C. sempervirens: nCup s 1 [10]; S. kali: nSal k 1 [28]; Chenopodium album: rChe a 1 [29], rChe a 2 (profilin) [30] and rChe a 3 (polcalcin) [31]; P. acerifolia :rPla a 1 and nPla a 2 (BIAL-Aristegui, Bilbao, Spain). As pan-allergens, we used polcalcin: rChe a 3 from C. album pollen [31]; nonspecific LTP from peach: rPru p 3 [32]; a mixture of 3 profilins: rCuc m 2 from muskmelon [33], rChe a 2 from C. album pollen [30] and rMal d 4 from apple [10].…”

“…As pan-allergens, we used polcalcin: rChe a 3 from C. album pollen [31]; nonspecific LTP from peach: rPru p 3 [32]; a mixture of 3 profilins: rCuc m 2 from muskmelon [33], rChe a 2 from C. album pollen [30] and rMal d 4 from apple [10]. We also tested purified proteins from wheat flour: nCM3 and nCM16 (α-amylase inhibitor subunits) [34]; lentil: nLen c 1 (vicilin) [35]; a mixture of 2 chitinases: n Pers a 1 from avocado [36] and rCas s 5 from chestnut [36]; a mixture of PR-10 protein: nBet v1 from B. verrucosa [37] and rMal d 1 from apple [38]; nSin a 1, a 2S albumin from mustard [39]; peroxidase as a CCD marker (Sigma-Aldrich Co. St. Louis, Mo., USA).…”

The Principle of Homologous Groups in Regulatory Affairs of Allergen Products – A Proposal

“…However, a homologous Ca 2+ -binding protein has been characterized in alder (Aln g 4). Additionally, the amino acid sequence identity of Bet v 4 is similar to several other Ca 2+ -binding proteins from botanically distant pollen allergens such as Phl p 7 from timothy grass [16, 17]. Its cross-reactivity with Phl p 7 has been demonstrated [18, 19] and can be explained by similar three-dimensional structure of both allergens [20].…”