title: Development of a strategy for the total chemical synthesis of an allergenic protein: the peach LTP Pru p 3.
abstract: The possibility to obtain allergenic proteins by means of total chemical synthesis would be a big step forward in the development of cures to food allergy and in the study of the mechanism of allergic reactions, because this would allow to achieve control at the molecular level over the structure of the product and to study its relationship with the allergenic activity in fine details. This is instead not possible by using allergens produced by extraction from natural sources or by recombinant DNA techniques. In this work, we aimed to test for the first time the feasibility of the total chemical synthesis of an allergenic protein. Pru p 3, the most studied member of the family of lipid transfer proteins, relevant plant food pan-allergens, was used as model target. Strategies for the convergent assembly of the target protein, starting from five peptide fragments to be bound by means of either native chemical ligation or peptide hydrazide ligation, followed by desulfurization, to achieve ligations at alanine, were developed and tested. All the reaction conditions were set up and optimized. Two large peptides covering the two halves of the protein sequence were synthesized and structurally characterized by means of circular dichroism, and their immunogenicity was proved by means of immunoblot, using antibodies against Pru p 3, and immunoCAP inhibition tests. Finally, the five peptides were bound together to produce the whole protein stretch. The obtained results demonstrate the feasibility of total chemical synthesis as a new way to obtain pure allergens. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.
authors.affiliation: Food and Drug Department, University of Parma, Viale delle Scienze 17/A, 43124, Parma, Italy.
authors.forename: Sofie
authors.initials: S
authors.lastname: Buhler
authors.affiliation: Department of Experimental Immunology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Jaap H
authors.initials: JH
authors.lastname: Akkerdaas
authors.affiliation: Transfusion Medicine Unit, ASMN-IRCCS, Reggio Emilia, Italy.|Department of Medicine and Surgery, Via Gramsci 14, 43125, Parma.
authors.forename: Thelma
authors.initials: T
authors.lastname: A Pertinhez
authors.affiliation: Department of Experimental Immunology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Ronald
authors.initials: R
authors.lastname: Van Ree
authors.affiliation: Food and Drug Department, University of Parma, Viale delle Scienze 17/A, 43124, Parma, Italy.
authors.forename: Arnaldo
authors.initials: A
authors.lastname: Dossena
authors.affiliation: Food and Drug Department, University of Parma, Viale delle Scienze 17/A, 43124, Parma, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Sforza
authors.affiliation: Food and Drug Department, University of Parma, Viale delle Scienze 17/A, 43124, Parma, Italy.
authors.forename: Tullia
authors.identifier: http://orcid.org/0000-0002-1109-1249
authors.initials: T
authors.lastname: Tedeschi
mesh_terms: D000485:Allergens / Q000737:chemistry*; D002352:Carrier Proteins / Q000138:chemical synthesis* / Q000737:chemistry; D006801:Humans; D015394:Molecular Structure; D000068239:Prunus persica / Q000737:chemistry*
chemical_list: D000485:Allergens; D002352:Carrier Proteins; C030363:lipid transfer protein
keywords: chemical ligation; food allergy; immunogenicity; protein synthesis