title: Purification and structural stability of the peach allergens Pru p 1 and Pru p 3.
abstract: 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 degrees C. Thus, Pru p 1 was unfolded at pH 3 even at 25 degrees C but was able to refold after heating to 95 degrees 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.
authors.affiliation: Department of Pathophysiology, Medical University of Vienna, Vienna, Austria.
authors.forename: Sonja
authors.initials: S
authors.lastname: Gaier
authors.forename: Justin
authors.initials: J
authors.lastname: Marsh
authors.forename: Christina
authors.initials: C
authors.lastname: Oberhuber
authors.forename: Neil M
authors.initials: NM
authors.lastname: Rigby
authors.forename: Alison
authors.initials: A
authors.lastname: Lovegrove
authors.forename: Stefano
authors.initials: S
authors.lastname: Alessandri
authors.forename: Peter
authors.initials: P
authors.lastname: Briza
authors.forename: Christian
authors.initials: C
authors.lastname: Radauer
authors.forename: Laurian
authors.initials: L
authors.lastname: Zuidmeer
authors.forename: Ronald
authors.initials: R
authors.lastname: van Ree
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Hemmer
authors.forename: Ana I
authors.initials: AI
authors.lastname: Sancho
authors.forename: Clare
authors.initials: C
authors.lastname: Mills
authors.forename: Karin
authors.initials: K
authors.lastname: Hoffmann-Sommergruber
authors.forename: Peter R
authors.initials: PR
authors.lastname: Shewry
mesh_terms: D000485:Allergens / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D052179:Antigens, Plant; D002942:Circular Dichroism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D010940:Plant Proteins; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D010940:Plant Proteins; C403252:Pru p 1 allergen; C463181:Pru p 3 allergen; D011994:Recombinant Proteins