title: Ca2+-binding allergens from olive pollen exhibit biochemical and immunological activity when expressed in stable transgenic Arabidopsis.
abstract: Employing transgenic plants as alternative systems to the conventional Escherichia coli, Pichia pastoris or baculovirus hosts to produce recombinant allergens may offer the possibility of having available edible vaccines in the near future. In this study, two EF-hand-type Ca2+-binding allergens from olive pollen, Ole e 3 and Ole e 8, were produced in transgenic Arabidopsis thaliana plants. The corresponding cDNAs, under the control of the constitutive CaMV 35S promoter, were stably incorporated into the Arabidopsis genome and encoded recombinant proteins, AtOle e 3 and AtOle e 8, which exhibited the molecular properties (i.e. MS analyses and CD spectra) of their olive and/or E. coli counterparts. Calcium-binding assays, which were carried out to assess the biochemical activity of AtOle e 3 and AtOle e 8, gave positive results. In addition, their mobilities on SDS/PAGE were according to the conformational changes derived from their Ca2+-binding capability. The immunological behaviour of Arabidopsis-expressed proteins was equivalent to that of the natural- and/or E. coli-derived allergens, as shown by their ability to bind allergen-specific rabbit IgG antiserum and IgE from sensitized patients. These results indicate that transgenic plants constitute a valid alternative to obtain allergens with structural and immunological integrity not only for scaling up production, but also to develop new kind of vaccines for human utilization.
authors.affiliation: Dpto. Bioquímica y Biología Molecular I, Universidad Complutense, Madrid, Spain.
authors.forename: Amalia
authors.initials: A
authors.lastname: Ledesma
authors.forename: Verónica
authors.initials: V
authors.lastname: Moral
authors.forename: Mayte
authors.initials: M
authors.lastname: Villalba
authors.forename: Julio
authors.initials: J
authors.lastname: Salinas
authors.forename: Rosalía
authors.initials: R
authors.lastname: Rodríguez
mesh_terms: D000485:Allergens / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D017360:Arabidopsis / Q000235:genetics*; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D031658:Olea / Q000276:immunology*; D010940:Plant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D030821:Plants, Genetically Modified; D011058:Pollen / Q000276:immunology*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D002135:Calcium-Binding Proteins; C102722:OLE3 protein, Olea europaea; C464977:Ole e 8 protein, olive; D010940:Plant Proteins; D011994:Recombinant Proteins; D002118:Calcium