Purification and structural stability of the peach allergens Pru p 1 and Pru p 3

Pru p 1 (a Bet v 1 homologue) and Pru p 3 (a nonspecific lipid transfer protein; nsLTP) are major allergenic proteins in peach fruit, but differ in their abundance and stability. Pru p 1 has low abundance and is highly labile and was purified after expression as a recombinant protein in Escherichia...

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Bibliographic Details
Published in:Molecular nutrition & food research 2008-11, Vol.52 (S2), p.S220-S229
Main Authors: Gaier, Sonja, Marsh, Justin, Oberhuber, Christina, Rigby, Neil M, Lovegrove, Alison, Alessandri, Stefano, Briza, Peter, Radauer, Christian, Zuidmeer, Laurian, van Ree, Ronald, Hemmer, Wolfgang, Sancho, Ana I, Mills, Clare, Hoffmann-Sommergruber, Karin, Shewry, Peter R
Format: Article
Language:English
Subjects:
Allergens - chemistry
Allergens - immunology
Allergens - isolation & purification
Antigens, Plant
Bet v 1 homologue
Circular Dichroism
Escherichia coli
Food allergy
Humans
Hydrogen-Ion Concentration
Lipid transfer protein
Magnetic Resonance Spectroscopy
Malus
Peach
Plant Proteins
Protein Structure, Secondary
Pru p 3
Prunus
Recombinant Proteins - chemistry
Recombinant Proteins - isolation & purification
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Summary:Pru p 1 (a Bet v 1 homologue) and Pru p 3 (a nonspecific lipid transfer protein; nsLTP) are major allergenic proteins in peach fruit, but differ in their abundance and stability. Pru p 1 has low abundance and is highly labile and was purified after expression as a recombinant protein in Escherichia coli. Pru p 3 is highly abundant in peach peel and was purified by conventional methods. The identities of the proteins were confirmed by sequence analysis and their masses determined by MS analysis. The purified proteins reacted with antisera against related allergens from other species: Pru p 1 with antiserum to Bet v 1 and Pru p 3 with antiserum to Mal d 3 (from apple). The presence of secondary and tertiary structure was demonstrated by circular dichroism (CD) and high field NMR spectroscopy. CD spectroscopy also showed that the two proteins differed in their stability at pH 3 and in their ability to refold after heating to 95°C. Thus, Pru p 1 was unfolded at pH 3 even at 25°C but was able to refold after heating to 95°C at pH 7.5. In contrast, Pru p 3 was unable to refold after heating under neutral conditions but readily refolded after heating at pH 3.
ISSN:1613-4125
1613-4133
1521-3803
DOI:10.1002/mnfr.200700274