Lipid transfer proteins from fruit: cloning, expression and quantification
| Autor: | Laurian, Zuidmeer, W Astrid, van Leeuwen, Ilona Kleine, Budde, Jessica, Cornelissen, Ingrid, Bulder, Ilona, Rafalska, Noèlia Telléz, Besolí, Jaap H, Akkerdaas, Riccardo, Asero, Montserrat, Fernandez Rivas, Montserrat Fernandez, Rivas, Eloina, Gonzalez Mancebo, Eloina Gonzalez, Mancebo, Ronald, van Ree |
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| Přispěvatelé: | Amsterdam institute for Infection and Immunity, Experimental Immunology, Other departments |
| Jazyk: | angličtina |
| Rok vydání: | 2005 |
| Předmět: |
Cloning
Organism Immunology Molecular Sequence Data Clone (cell biology) Biology law.invention Mice law Protein biosynthesis Immunology and Allergy Animals Amino Acid Sequence Peptide sequence Plant Proteins Cloning Nucleic acid sequence General Medicine Allergens Antigens Plant Biochemistry Fruit Malus Protein Biosynthesis Recombinant DNA Prunus Rabbits Carrier Proteins Plant lipid transfer proteins |
| Zdroj: | International archives of allergy and immunology, 137(4), 273-281. S. Karger AG |
| ISSN: | 1018-2438 |
| Popis: | Background: Lipid transfer proteins (LTP) are stable, potentially life-threatening allergens in fruits and many other vegetable foods. The aim of this study was to clone and express recombinant apple LTP (Mal d 3), as has previously been done for peach LTP (Pru p 3) and set up quantitative tests for measuring fruit LTPs. Methods: cDNA for Mal d 3 and Pru p 3 was cloned, expressed in the yeast Pichia pastoris and the resulting proteins were purified via cation exchange chromatography. The immune reactivity of rMal d 3 was compared to nMal d 3 by RAST (inhibition), immunoblotting and basophil histamine release testing. To obtain monoclonal and monospecific polyclonal antibodies, mice and rabbits were immunized with purified nMal d 3. Results: The deduced amino acid sequence of Mal d 3 was identical to the published sequence, Pru p 3 differed at two positions (S9A and S76H). The rMal d 3 had an IgE-binding potency and biological activity close to its natural counterpart. One sandwich ELISA selectively detecting apple LTP and another cross-reactive with cherry, nectarine and hazelnut LTP were developed. In addition, a competitive RIA was developed with polyclonal rabbit antiserum and labeled nMal d 3. Conclusion: rMal d 3 (as shown before for rPru p 3) may be a useful tool for application in component-resolved diagnosis of food allergy. Assays for the measurement of LTP will increase the traceability of this potentially dangerous allergen. |
| Databáze: | OpenAIRE |
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