Mapping of Malus domestica allergens by 2-D electrophoresis and IgE-reactivity.
The importance of apple allergens has been repeatedly emphasized, and their presence has been confirmed both in pollen and in fruits. In the present study, a combination of proteomic tools have been used to build a complete allergen map of apple. The water-soluble fraction of an apple extract was precipitated using a phenol-based procedure and separated by 2-DE. Initially four previously classified allergens, Mal d 1, Mal d 2, Mal d 3 and Mal d 4, could be identified in Western blots with polyclonal rabbit antibodies directed to the four respective allergens, and subsequently matched to the bands recognized by several patient sera. Further, all four known apple allergens were localized on a 2-DE map and they were matched with spots recognized by sera of patients with different allergic patterns. Moreover, a new, putative allergen could be identified using MS. We evaluated the influence of post-translational modifications and the immunoreactivity under different analytical conditions. The comparison of different visualization methods for 2-DE gels and blots revealed that even very low concentrations of the intact epitopes are detectable by IgEs of patients, and therefore might be sufficient to trigger allergic symptoms in sensitized individuals.
Herndl
Anita
A
Plant Biotechnology Unit, Department of Biotechnology, BOKU, Vienna, Austria.
Marzban
Gorji
G
Kolarich
Daniel
D
Hahn
Rainer
R
Boscia
Donato
D
Hemmer
Wolfgang
W
Maghuly
Fatemeh
F
Stoyanova
Elitza
E
Katinger
Hermann
H
Laimer
Margit
M
D000485:Allergens / Q000032:analysis / Q000276:immunology*; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007106:Immune Sera / Q000032:analysis; D007073:Immunoglobulin E / Q000276:immunology*; D027845:Malus / Q000737:chemistry*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000032:analysis / Q000276:immunology; D020543:Proteome / Q000032:analysis / Q000276:immunology*; D021241:Spectrometry, Mass, Electrospray Ionization
D000485:Allergens; D052179:Antigens, Plant; D007106:Immune Sera; C095551:MALD1 protein, Malus domestica; D010940:Plant Proteins; D020543:Proteome; D007073:Immunoglobulin E