Molecular Characterization of Recombinant Mus a 5 Allergen from Banana Fruit
- 435 Downloads
Allergy to banana fruit appears to have become an important cause of fruit allergy in Europe. Among five allergens that have been found, beta-1,3-glucanase denoted as Mus a 5 was identified as a candidate allergen for the component-resolved allergy diagnosis of banana allergy. Because of the variations in protein levels in banana fruit, in this study Mus a 5 was produced as a fusion protein with glutathione-S-transferase in Escherichia coli. The recombinant Mus a 5 was purified under native conditions by a combination of affinity, ion-exchange, and reversed phase chromatography. N-terminal sequence was confirmed by Edman degradation and 55 % of the primary structure was identified by mass fingerprint, while the secondary structure was assessed by circular dichroism spectroscopy. IgG reactivity of recombinant protein was shown in 2-D immunoblot with anti-Mus a 5 antibodies, while IgG and IgE binding to natural Mus a 5 was inhibited with the recombinant Mus a 5 in immunoblot inhibition test. IgE reactivity of recombinant Mus a 5 was shown in ELISA within a group of ten persons sensitized to banana fruit. Recombinant Mus a 5 is a novel reagent suitable for the component-resolved allergy diagnosis of banana allergy.
KeywordsAllergen Banana glucanase Food allergy IgE reactivity Mus a 5
This study was supported by the Ministry of Education, Science and Technological Development, Republic of Serbia, Grant No. 172049. The authors acknowledge support of the FP7 RegPot project FCUB ERA GA No. 256716. The EC does not share responsibility for the content of the article.
- 5.Ciardiello, M. A., Giangrieco, I., Tuppo, L., Tamburrini, M., Buccheri, M., Palazzo, M. L., et al. (2009). Influence of the natural ripening stage, cold storage, and ethylene treatment on the protein and IgE-bunding profiles of Green and Gold kiwi fruit extracts. Journal Agricultural of Food Chemistry, 57(4), 1565–1571.CrossRefGoogle Scholar
- 7.Peumans, W., Proost, P., Swennen, R., & Van Damme, E. (2002). The abundant class III chitinase homolog in young developing banana fruits behaves as a transient vegetative storage protein and most probably serves as an important supply of amino acids for the synthesis of ripening-associated proteins. Plant Physiology, 130(2), 1063–1072.CrossRefGoogle Scholar
- 8.Choudhury, S. R., Roy, S., Singh, S. K., & Sengupta, D. N. (2010). Molecular characterization and differential expression of beta-1,3-glucanase during ripening in banana fruit in response to ethylene, auxin, ABA, wounding, cold and light-dark cycles. Plant Cell Reports, 29(8), 813–828.CrossRefGoogle Scholar
- 13.Palomares, O., Villalba, M., & Rodriguez, R. (2003). The C-terminal segment of the 1,3-beta-glucanase Ole e 9 from olive (Olea europeae) pollen is an independent domain with allergenic activity: Expression in Pichia pastoris and characterization. Biochemistry Journal, 369, 593–601.CrossRefGoogle Scholar
- 16.Gavrovic-Jankulovic, M., Poulsen, K., Brckalo, T., Bobic, S., Lindner, B., & Petersen, A. (2008). A novel recombinantly produced banana lectin isoform is a valuable tool for glycoproteomics and a potent modulator of the proliferation response in CD3+, CD4+, and CD8+ populations of human PBMCs. International Journal of Biochemistry Cell Biology, 40(5), 929–941.CrossRefGoogle Scholar
- 21.Harlow, E., & Lane, D. (1988). Antibodies: A laboratory manual. New York: Cold Spring Harbor Laboratory. 140.Google Scholar
- 22.MacGregor, E. A., & Balance, G. M. (1991). Possible secondary structure in plant and yeast β-glucanase. Biochemical Journal, 274, 41–43.Google Scholar
- 25.Lorenz, A.R., Haustein, D., & Vieths, S. (2001). IgE-binding propertis of recombinant food allergens. In Internet Symposium on Food Allergens. http://www.food-allergens.de/symposium-3-1/lorenz/lorenz-frame.html. 3(1), 1–36.
- 27.Teuber, S. S., Jarvis, K. C., Dandekar, A. M., Peterson, R. W., & Ansari, A. A. (1999). Identification and cloning of a complementary DNA encoding a vicilin-like proprotein, Jug r 2, from English walnut kernel (Juglans regia), a major food allergen. Journal of Allergy and Clinical Immunology, 104, 1311–1320.CrossRefGoogle Scholar
- 28.Barre, A., Culerrier, R., Granier, C., Selman, L., Peumans, W. J., Van Damme, E. J. M., et al. (2009). Mapping of IgE-binding epitopes on the major latex allergen Hev b 2 and the cross-reacting 1,3-beta-glucanase fruit allergens as a molecular basis for the latex-fruit syndrome. Molecular Immunology, 46, 1595–1604.CrossRefGoogle Scholar
- 29.Choudhury, S. R., Roy, S., & Sengupta, D. N. (2009). Characterization of cultivar differences in beta-1,3 glucanase gene expression, glucanase activity and fruit pulp softening rates during fruit ripening in three naturally occurring banana cultivars. Plant Cell Reports, 28(11), 1641–1653.CrossRefGoogle Scholar