Rodríguez García, Rosalía

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First Name
Last Name
Rodríguez García
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Biológicas
Bioquímica y Biología Molecular
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Now showing 1 - 6 of 6
  • Publication
    Characterization of Profilin and Polcalcin Panallergens From Ash Pollen
    (Esmon Publicidad, 2014) Mas García, Salvador; Garrido Arandia, María; Batanero Cremades, Eva; Purohit, A.; Pauli, G.; Rodríguez García, Rosalía; Barderas Manchado, Rodrigo; Villalba Díaz, Mayte
    Background: Ash (Fraxinus excelsior) is an important source of allergenic pollen in temperate areas of Europe. Profilin and polcalcin are 2 important panallergens involved in cross-reactivity between different sources. Objective: To clone and produce Fra e 2 (profilin) and Fra e 3 (polcalcin) as recombinant proteins and evaluate their immunological properties using the natural forms obtained from ash pollen. Methods: Total RNA from ash pollen was used as a template to obtain the specific complementary DNA (cDNA) sequences of the 2 panallergens. The cDNA-encoding sequences were cloned into the pET11b expression vector and used to transform BL21 (DE3) Escherichia coli cells. Proteins were expressed, purified by chromatography, and characterized structurally by circular dichroism, mass spectrometry, and immunologically by western blot and ELISA using profilin and polcalcin polyclonal antibodies and human sera from ash pollen–sensitized patients. Results: Profilin and polcalcin amino acid sequences from ash pollen showed a high degree of identity with homologous allergens from different sources. The cDNA-encoding allergen sequences were expressed as nonfusion recombinant proteins and purified to homogeneity. Secondary structure values were similar to those obtained from other members of these families. Allergenic Properties of the recombinant allergens were observed to be equivalent to those of the natural counterparts of F excelsior pollen.Conclusions: Fra e 2 and Fra e 3 recombinant allergens might be used in clinical diagnosis to determine profilin- and polcalcin-specific IgE levels present in the sera of ash pollen–sensitized patients, thus facilitating the finding of the sensitizing source in areas with complex sensitization profiles.
  • Publication
    Pollensomes as natural vehicles for pollen allergens
    (American Association of Immunologists, 2015-07-15) Prado, Noela; De Linares, C.; Sanz, M.L.; Gamboa, P.; Villalba Díaz, Mayte; Rodríguez García, Rosalía; Batanero Cremades, Eva
    Olive (Olea europaea) pollen constitutes one of the most important allergen sources in the Mediterranean countries and some areas of the United States, South Africa, and Australia. Recently, we provided evidence that olive pollen releases nanovesicles of respirable size, named generically pollensomes, during in vitro germination. Olive pollensomes contain allergens, such as Ole e 1, Ole e 11, and Ole e 12, suggesting a possible role in allergy. The aim of this study was to assess the contribution of pollensomes to the allergic reaction. We show that pollensomes exhibit allergenic activity in terms of patients' IgE-binding capacity, human basophil activation, and positive skin reaction in sensitized patients. Furthermore, allergen-containing pollensomes have been isolated from three clinically relevant nonphylogenetically related species: birch (Betula verrucosa), pine (Pinus sylvestris), and ryegrass (Lolium perenne). Most interesting, pollensomes were isolated from aerobiological samples collected with an eight-stage cascade impactor collector, indicating that pollensomes secretion is a naturally occurring phenomenon. Our findings indicate that pollensomes may represent widespread vehicles for pollen allergens, with potential implications in the allergic reaction.
  • Publication
    Amaranthaceae Pollens: Review of an Emerging Allergy in the Mediterranean Area
    (Esmon Publicidad, 2014) Villalba, Mayte; Barderas, R.; Mas, S.; Colás, S.; Batanero, E.; Rodríguez García, Rosalía
    The Amaranthaceae family is composed of about 180 genera and 2500 species. These common weeds have become increasingly relevant as triggers of allergy in the last few years, as they are able to rapidly colonize salty and arid soils in extensive desert areas. The genera Chenopodium, Salsola, and Amaranthus are the major sources of pollinosis from the Amaranthaceae family in southern Europe, western United States, and semidesert areas of Saudi Arabia, Kuwait, and Iran. In Spain, Salsola kali is one of the most relevant causes of pollinosis, together with olive and grasses. To date, 9 Amaranthaceae pollen allergens from Chenopodium album, Salsola kali, and Amaranthus retroflexus have been described and are listed in the International Union of Immunological Societies allergen nomenclature database. The major allergens of Amaranthaceae pollen belong to the pectin methylesterase, Ole e 1–like, and profilin panallergen families, whereas the minor allergens belong to the cobalaminindependent methionine synthase and polcalcin panallergen families. These relevant allergens have been characterized physicochemically, and immunologically at different levels. Recombinant forms, allergenic fusion recombinant proteins, and hypoallergenic derivatives of these allergens have been expressed in bacteria and yeast and compared with their natural proteins from pollen. In this review, we provide an extensive overview of Amaranthaceae pollen allergens, focusing on their physicochemical, and immunological properties and on their clinical significance in allergic patients. We also review studies where these recombinant allergens and their hypoallergenic derivatives have been used in clinical diagnosis and their potential use in personalized therapy.
  • Publication
    Ash pollen immunoproteomics: Identification, immunologic characterization, and sequencing of 6 new allergens
    (Elsevier, 2014) Mas García, Salvador; Torres Pardo, María; Garrido-Arandia, María; Salamanca, Guillermo; Castro, Lourdes; Barral, Patricia; Purohit, Ashok; Pauli, Gabrielle; Rodríguez García, Rosalía; Batanero Cremades, Eva; Barderas Manchado, Rodrigo; Villalba Díaz, Mayte
    Immunoproteomics, IgE-inhibition assays and cDNA-cloning reveals that ash and olive allergenic protein profiles are mostly equivalent, thus explaining their high cross reactivity. Our data suggest simplifying diagnosis of patients by using indistinctly ash or olive pollen.
  • Publication
    An Enzymatically Active β-1,3-Glucanase from Ash Pollen with Allergenic Properties: A Particular Member in the Oleaceae Family
    (Plos, 2015-07) Torres, María; Palomares, Oscar; Quiralte, Joaquín; Pauli, Gabrielle; Rodríguez García, Rosalía; Villalba, Mayte
    Endo-β-1,3-glucanases are widespread enzymes with glycosyl hydrolitic activity involved in carbohydrate remodelling during the germination and pollen tube growth. Although members of this protein family with allergenic activity have been reported, their effective contribution to allergy is little known. In this work, we identified Fra e 9 as a novel allergenic β-1,3-glucanase from ash pollen. We produced the catalytic and carbohydrate-binding domains as two independent recombinant proteins and characterized them from structural, biochemical and immunological point of view in comparison to their counterparts from olive pollen. We showed that despite having significant differences in biochemical activity Fra e 9 and Ole e 9 display similar IgE-binding capacity, suggesting that β-1,3-glucanases represent an heterogeneous family that could display intrinsic allergenic capacity. Specific cDNA encoding Fra e 9 was cloned and sequenced. The full-length cDNA encoded a polypeptide chain of 461 amino acids containing a signal peptide of 29 residues, leading to a mature protein of 47760.2 Da and a pI of 8.66. An N-terminal catalytic domain and a C-terminal carbohydrate-binding module are the components of this enzyme. Despite the phylogenetic proximity to the olive pollen β-1,3-glucanase, Ole e 9, there is only a 39% identity between both sequences. The N- and C-terminal domains have been produced as independent recombinant proteins in Escherichia coli and Pichia pastoris, respectively. Although a low or null enzymatic activity has been associated to long β-1,3-glucanases, the recombinant N-terminal domain has 200-fold higher hydrolytic activity on laminarin than reported for Ole e 9. The C-terminal domain of Fra e 9, a cysteine-rich compact structure, is able to bind laminarin. Both molecules retain comparable IgE-binding capacity when assayed with allergic sera. In summary, the structural and functional comparison between these two closely phylogenetic related enzymes provides novel insights into the complexity of β-1,3-glucanases, representing a heterogeneous protein family with intrinsic allergenic capacity.
  • Publication
    The Allergenic Structure of the Thaumatinlike Protein Ole e 13 Is Degraded by Processing of Raw Olive Fruits
    (Esmon Publicidad, 2014) Torres, M.; Alvarez García, Elvira; Bartra, J.; Alcántara, M.; Palomares Gracia, Oscar; Villalba Díaz, Mayte; Rodríguez García, Rosalía
    Background: The thaumatin-like protein (TLP) Ole e 13 in raw olive fruit is responsible for occupational allergy in olive oil mill workers. However,these workers do not experience allergic symptoms after ingestion of edible olive.Objectives: To analyze the presence of IgE-reactive TLP in raw and edible olive fruit and to assess the allergenic potency of both sources.Methods: The content of TLP in raw and edible olive fruit protein extracts was analyzed using immunoblotting with sera from allergic patients and with olive TLP–specific IgG. The structural and immunological stability of TLP were assayed using immunoblotting after treatment of both raw olive and purified TLP with 0.25 M NaOH solution for 24 hours. Olive pollen extract was investigated by immunoblotting for TLP content. Results: The TLP contained in raw olive fruit was not present in edible olives as a result of maceration before human consumption. No TLP was detected in olive pollen using specific IgG or sera from patients allergic to olive fruit. Sera from patients allergic to olive pollen did not react with purified TLP. Conclusions: IgE-reactive TLP is not present in edible olive, thus explaining the low number of patients allergic to this highly consumed fruit. Patients allergic to olive pollen are not sensitized to TLP and, therefore, not expected to be at risk of food allergy to olive fruit or TLP plant sources