Amb a 1 isoforms: Unequal siblings with distinct immunological features.

BACKGROUND: Ragweed pollen represents a major allergy risk factor. Ragweed extracts contain five different isoforms of the major allergen Amb a 1. However, the immunological characteristics of Amb a 1 isoforms are not fully investigated. Here, we compared the physicochemical and immunological properties of three most important Amb a 1 isoforms. METHODS: After purification, the isoforms were physicochemically characterized, tested for antibody binding and induction of human T-cell proliferative responses. Their immunological properties were further evaluated in vitro and in vivo in a mouse model. RESULTS: Amb a 1 isoforms exhibited distinct patterns of IgE binding and immunogenicity. Compared to Amb a 1.02 or 03 isoforms, Amb a 1.01 showed higher IgE-binding activity. Isoforms 01 and 03 were the most potent stimulators of patients' T cells. In a mouse model of immunization, Amb a 1.01 induced higher levels of IgG and IgE antibodies when compared to isoforms 02 and 03. Interestingly, ragweed-sensitized patients also displayed an IgG response to Amb a 1 isoforms. However, unlike therapy-induced antibodies, sensitization-induced IgG did not show IgE-blocking activity. CONCLUSION: The present study showed that naturally occurring isoforms of Amb a 1 possess different immunogenic and sensitizing properties. These findings should be considered when selecting sequences for molecule-based diagnosis and therapy for ragweed allergy. Due to its high IgE-binding activity, isoform Amb a 1.01 should be included in diagnostic tests. In contrast, due to their limited B- and T-cell cross-reactivity patterns, a combination of different isoforms might be a more attractive strategy for ragweed immunotherapy.

Tree pollen allergens-an update from a molecular perspective.

It is estimated that pollen allergies affect approximately 40% of allergic individuals. In general, tree pollen allergies are mainly elicited by allergenic trees belonging to the orders Fagales, Lamiales, Proteales, and Pinales. Over 25 years ago, the gene encoding the major birch pollen allergen Bet v 1 was the first such gene to be cloned and its product characterized. Since that time, 53 tree pollen allergens have been identified and acknowledged by the WHO/IUIS allergen nomenclature subcommittee. Molecule-based profiling of allergic sensitization has helped to elucidate the immunological connections of allergen cross-reactivity, whereas advances in biochemistry have revealed structural and functional aspects of allergenic proteins. In this review, we provide a comprehensive overview of the present knowledge of the molecular aspects of tree pollen allergens. We analyze the geographic distribution of allergenic trees, discuss factors pivotal for allergic sensitization, and describe the role of tree pollen panallergens. Novel allergenic tree species as well as tree pollen allergens are continually being identified, making research in this field highly competitive and instrumental for clinical applications.

Bet v 1--a Trojan horse for small ligands boosting allergic sensitization?

BACKGROUND: Birch pollen allergy represents the main cause of winter and spring pollinosis in the temperate climate zone of the northern hemisphere and sensitization towards Bet v 1, the major birch pollen allergen, affects over 100 million allergic patients. The major birch pollen allergen Bet v 1 has been described as promiscuous acceptor for a wide variety of hydrophobic ligands. OBJECTIVE: In search of intrinsic properties of Bet v 1, which account responsible for the high allergenic potential of the protein, we thought to investigate the effects of ligand-binding on immunogenic as well as allergenic properties. METHODS: As surrogate ligand of Bet v 1 sodium deoxycholate (DOC) was selected. Recombinant and natural Bet v 1 were characterised physico-chemically as well as immunologically in the presence or absence of DOC, and an animal model of allergic sensitization was established. Moreover, human IgE binding to Bet v 1 was analysed by nuclear magnetic resonance (NMR) spectroscopy. RESULTS: Ligand-binding had an overall stabilizing effect on Bet v 1. This translated in a Th2 skewing of the immune response in a mouse model. Analyses of human IgE binding on Bet v 1 in mediator release assays revealed that ligand-bound allergen-induced degranulation at lower concentrations; however, in basophil activation tests with human basophils ligand-binding did not show this effect. For the first time, human IgE epitopes on Bet v 1 were determined using antibodies isolated from patients' sera. The IgE epitope mapping of Bet v 1 demonstrated the presence of multiple binding regions. CONCLUSIONS AND CLINICAL RELEVANCE: Deoxycholate binding stabilizes conformational IgE epitopes on Bet v 1; however, the epitopes themselves remain unaltered. Therefore, we speculate that humans are exposed to both ligand-bound and free Bet v 1 during sensitization, disclosing the ligand-binding cavity of the allergen as key structural element.

Bet v 1-like pollen allergens of multiple Fagales species can sensitize atopic individuals.

BACKGROUND: In the temperate climate zone of the Northern hemisphere, Fagales pollen allergy represents the main cause of winter/spring pollinosis. Among Fagales trees, pollen allergies are strongly associated within the Betulaceae and the Fagaceae families. It is widely accepted that Fagales pollen allergies are initiated by sensitization against Bet v 1, the birch pollen major allergen, although evidence is accumulating that the allergenic activity of some Bet v 1-like molecules has been underestimated. OBJECTIVE: To investigate the allergenic potential of the clinically most important Fagales pollen allergens from birch, alder, hazel, hornbeam, hop-hornbeam, oak, beech and chestnut. METHODS: To obtain the full spectrum of allergens, the three previously unavailable members of the Bet v 1-family, hop-hornbeam Ost c 1, chestnut Cas s 1 and beech Fag s 1, were identified in the respective pollen extracts, cloned and produced as recombinant proteins in E. coli. Together with recombinant Bet v 1, Aln g 1, Car b 1, Cor a 1 and Que a 1, the molecules were characterized physicochemically, mediator release assays were performed and IgE cross-reactivity was evaluated by ELISA and Immuno Solid-phase Allergen Chip (ISAC) IgE inhibition assays. RESULTS: All allergens showed the typical Bet v 1-like secondary structure elements, and they were all able to bind serum IgE from Fagales allergic donors. Strong IgE binding was observed for Betuloideae and Coryloideae allergens, however, cross-reactivity between the two subfamilies was limited as explored by inhibition experiments. In contrast, IgE binding to members of the Fagaceae could be strongly inhibited by serum pre-incubation with allergens of the Betuloideae subfamily. CONCLUSIONS AND CLINICAL RELEVANCE: The data suggest that Bet v 1-like allergens of the Betuloideae and Coryloideae subfamily might have the potential to induce IgE antibodies with different specificities, while allergic reactions towards Fagaceae allergens are the result of IgE cross-reactivity.

Unbiased Quantitative Proteomics Reveals a Crucial Role of the Allergen Context for the Activation of Human Dendritic Cells.

Worldwide, more than 1 billion people suffer from allergic diseases. However, until now it is not fully understood how certain proteins can induce allergic immune responses, while others cannot. Studies suggest that allergenicity is a process not only determined by properties of the allergen itself but also by costimulatory factors, that are not classically associated with allergic reactions. To investigate the allergenicity of the major birch pollen allergen Bet v 1 and the impact of adjuvants associated with pollen, e.g. lipopolysaccharide (LPS), we performed quantitative proteome analysis to study the activation of monocyte-derived dendritic cells (moDCs). Thus, we treated cells with birch pollen extract (BPE), recombinant Bet v 1, and LPS followed by proteomic profiling via high-performance liquid chromatography and tandem mass spectrometry (HPLC-MS/MS) using isobaric labelling. Enrichment and pathway analysis revealed the influence of regulated proteins especially in cytokine signalling and dendritic cell activation. We found highly regulated, but differentially expressed proteins after treatment with BPE and LPS, whereas the cellular response to Bet v 1 was limited. Our findings lead to the conclusion that Bet v 1 needs a specific "allergen context" involving cofactors apart from LPS to induce an immune response in human moDCs.

1H, 13C and 15N resonance assignments and second structure information of Fag s 1: Fagales allergen from Fagus sylvatica.

Fagales allergens belonging to the Bet v 1 family account responsible for the majority of spring pollinosis in the temperate climate zones in the Northern hemisphere. Among them, Fag s 1 from beech pollen is an important trigger of Fagales pollen associated allergic reactions. The protein shares high similarity with birch pollen Bet v 1, the best-characterized member of this allergen family. Of note, recent work on Bet v 1 and its homologues found in Fagales pollen demonstrated that not all allergenic members of this family have the capacity to induce allergic sensitization. Fag s 1 was shown to bind pre-existing IgE antibodies most likely primarily directed against other members of this multi-allergen family. Therefore, it is especially interesting to compare the structures of Bet v 1-like pollen allergens, which have the potential to induce allergic sensitization with allergens that are mainly cross-reactive. This in the end will help to identify allergy eliciting molecular pattern on Bet v 1-like allergens. In this work, we report the (1)H, (15)N and (13)C NMR assignment of beech pollen Fag s 1 as well as the secondary structure information based on backbone chemical shifts.