A non-allergenic Ole e 1-like protein from birch pollen as a tool to design hypoallergenic vaccine candidates.
ABSTRACT Recombinant DNA technology offers several approaches to convert allergens into hypoallergenic derivatives that can represent the basis of novel, safer and more effective forms of allergy vaccines. In this context, we used a new strategy for the design of a hypoallergenic derivative of Ole e 1, the main allergen of olive pollen. By screening a cDNA library from birch pollen, the clone BB18, encoding the birch counterpart of Ole e 1, was identified. In this study, BB18 has been produce in Pichia pastoris as a recombinant protein and immunologically characterized. The well-established non-allergenic properties of BB18 were used to generate a genetic variant of Ole e 1, named OB(55-58), by site-direct mutagenesis of four residues (E(55)V(56)G(57)Y(58)) in an IgE/IgG epitope of Ole e 1 by the corresponding ones in BB18 (SDSE). OB(55-58) was expressed in P. pastoris, purified to homogeneity and analyzed for IgE-reactivity by means of ELISA using sera from olive pollen allergic patients and rat basophil activation assay. T cell reactivity was assayed in a mouse model of Ole e 1 sensitization. The mutant OB(55-58) exhibited an impaired IgE reactivity, but not affected T cell reactivity, compared to wild type rOle e 1. This study emphasizes the usefulness of BB18 as a tool for epitope mapping and for engineering hypoallergenic derivatives of Ole e 1 as vaccine candidates for allergy prevention and treatment.
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ABSTRACT: Olive tree is one of the main allergy sources in Mediterranean countries. The identification of the allergenic repertoire from olive pollen has been essential for the development of rational strategies of standardization, diagnosis, and immunotherapy, all of them focused to increase the life quality of the patients. From its complex allergogram, twelve allergens -Ole e 1 to Ole e 12- have been identified and characterized to date. Most of them have been cloned and produced as recombinant forms, whose availability have allowed analyzing their three-dimensional structures, mapping their T-cell and B-cell epitopes, and determining the precise allergenic profile of patients for a subsequent patient-tailored immunotherapy. Protein mutant, hypoallergenic derivatives, or recombinant fragments have been also useful experimental tools to analyze the immune recognition of allergens. To test these molecules before using them for clinic purposes, a mouse model of allergic sensitizations has been used. This model has been helpful for assaying different prophylactic approaches based on tolerance induction by intranasal administration of allergens or hypoallergens, used as free or integrated in different delivery systems, and their findings suggest a promising utilization as nasal vaccines. Exosomes -nanovesicles isolated from bronchoalveolar lavage fluid of tolerogenic mice- have shown immunomodulatory properties, being able to protect mice against sensitization to Ole e 1.Methods 08/2013; · 3.64 Impact Factor
- The Journal of allergy and clinical immunology 04/2014; · 12.05 Impact Factor
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ABSTRACT: Since the 1970s, the establishment and development of the biotech industry has improved exponentially, allowing the commercial production of biopharmaceutical proteins. Nowadays, new recombinant protein production is considered a multibillion-dollar market, in which about 25% of commercial pharmaceuticals are biopharmaceuticals. But to achieve a competitive production process is not an easy task. Any production process has to be highly productive, efficient and economic. Despite that the perfect host is still not discovered, several research groups have chosen Pichia pastoris as expression system for the production of their protein because of its many features. The attempt of this review is to embrace several research lines that have adopted Pichia pastoris as their expression system to produce a protein on an industrial scale in the health care industry.Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology]. 12/2013; 44(4):1043-1048.