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Notes: Lipid-transfer proteins (LTPs), but not Bet v 1 homologues, have been identified as major allergens of apple and peach in the Rosaceae fruit-allergic population in the Mediterranean area. Many of these patients show cosensitization to mugwort pollen. LTPs have an ubiquitous distribution in tissues of many plant species, and have been proposed as a novel type of plant panallergens.〈section xml:id="abs1-2"〉〈title type="main"〉ObjectiveWe sought to isolate LTPs from Artemisia pollen and from a plant food not belonging to the Rosaceae family, such as chestnut nut, and to compare their amino acid sequences and IgE-binding capacities with those of apple and peach LTPs.〈section xml:id="abs1-3"〉〈title type="main"〉MethodsAllergens (LTPs) were isolated by different chromatographic methods (gel-filtration, ion exchange and/or reverse-phase HPLC), and characterized by N-terminal amino acid sequencing and MALDI analysis. Specific IgE-quantification and immunodetection, as well as immunoblot and ELISA inhibition assays, were carried out using sera from patients allergic to both apple and peach.〈section xml:id="abs1-4"〉〈title type="main"〉ResultsPurified LTPs from Artemisia pollen and from chestnut seed showed molecular masses about 9 700d, and 43–50% sequence identity with the equivalent allergens of apple and peach in the first 30 N-terminal residues, which comprise about one third of the total amino acid sequence. A similar degree of sequence identity (50%) was found between the Artemisia and chestnut proteins. Both isolated LTPs bound specific IgE of sera from Rosaceae fruits allergic patients. However, substantially lower values of specific IgE-binding and maximum ELISA inhibition percentages were obtained for Artemisia and chestnut LTPs when compared to those from apple and peach.〈section xml:id="abs1-5"〉〈title type="main"〉ConclusionLTPs from Artemisia pollen and chestnut crossreact with allergens (LTPs) of Rosaceae fruits, but significant differences in specific IgE-binding capacities were observed among members of the plant LTP family. Thus, further studies are needed to evaluate the clinical significance of the observed cross-reactivities of plant LTPs.

Notes: Background Artemisia vulgaris is a widespread weed in the Mediterranean area and several allergens have been detected in its pollen. One of them, Art v 3, belongs to the lipid-transfer protein (LTP) family and its prevalence in Artemisia-sensitized patients or its relationship with other LTP allergens is not clear.Objective To assess the pattern of sensitization to an array of mugwort allergens in a Mediterranean population, and to study the cross-reactivity of Art v 3 with Pru p 3 and Par j 1, relevant LTP allergens in the area.Methods Skin prick test was performed with whole extracts (A. vulgaris, Parietaria judaica and peach) and pure natural allergens Art v 1, Art v 3, Art v 60 kDa and Par j 1 in 24 mugwort-allergic patients from a Mediterranean area. In vitro assays included measurement of specific IgE and ELISA inhibition among LTP allergens.Results The three Artemisia allergens elicited a positive skin response in 70–80% of the patients. Seven patients were clearly sensitized to Par j 1 and 11 to Pru p 3. There was no correlation between Par j 1 and Pru p 3 sensitization, but a highly significant correlation was found between peach extract and Art v 3 as regards the skin response. No IgE cross-reactivity was observed between Art v 3/Par j 1 or Pru p 3/Par j 1. In contrast, Art v 3 significantly inhibited the binding to Pru p 3 of IgE from three patients' sera out of six studied, but Pru p 3 was not able to inhibit the IgE binding to Art v 3.Conclusion Art v 3 is a major mugwort allergen and in some patients with IgE to both Art v 3 and Pru p 3, Art v 3 behaves as the primary sensitizing agent.

Notes: Background Lipid-transfer proteins (LTPs) have been identified as major allergens of Rosaceae fruits in populations living in areas virtually free of Fagales trees, such as several Mediterranean communities. Pru p 3 and Mal d 3, the allergens from peach and apple, respectively, have a main clinical relevance in these areas.Objetive To isolate and characterize cDNAs for Pru p 3 and Mal d 3,and to produce recombinant Pru p 3 in the yeast Pichia pastoris.Methods cDNAs for both allergens were isolated by polymerase chain reaction using non-degenerated primers. Expression of Pru p 3 was performed in P. pastoris using the pPIC 9 vector. The recombinant product was purified by gel-filtration chromatography followed by RP-HPLC. Immunodetection and immunoblot inhibition assays were carried out with sera from peach-allergic patients.Results The cDNAs for both Pru p 3 and Mal d 3 showed a 273 open reading frame coding for the 91 amino acid mature polypeptides. The deduced amino acid sequences exhibited N-terminal regions fully identical to those previously determined for the natural peach and apple allergens. Pru p 3 was expressed in P. pastoris at 20 mg/L of culture medium. The recombinant allergen showed the same N-terminal sequence (plus a glutamic acid added for proper extracellular expression) and apparent molecular size as natural Pru p 3. Both the recombinant and natural forms of Pru p 3 displayed similar immunoglobulin (Ig)E-binding capacity in immunodetection and immunoblot inhibition assays.Conclusions Comparison of the complete primary structures of mature Pru p 3 and Mal d 3 deduced from their corresponding cDNA clones supports the close relationship between both allergens. Recombinant Pru p 3 binds IgE in vitro like its natural counterpart. Therefore, it can be a useful tool for specific diagnosis and structural studies.