Heat-induced conformational changes of Ara h 1, a major peanut allergen, do not affect its allergenic properties.
Ara h 1, a major peanut allergen was isolated, and its structure on secondary, tertiary, and quaternary level at ambient temperature was investigated using spectroscopic and biochemical techniques. Ara h 1 appeared to be a highly structured protein on a secondary level, possesses a clear tertiary fold, and is present as a trimeric complex. Heat treatment of purified Ara h 1 results in an endothermic, irreversible transition between 80 and 90 degreesC, leading to an increase in beta-structures and a concomitant aggregation of the protein. Ara h 1 from peanuts that were heat-treated prior to the purification procedure exhibited a similar denatured state with an increased secondary folding and a decreased solubility. The effect of heat treatment on the in vitro allergenic properties of Ara h 1 was investigated by means of a fluid-phase IgE binding assay using serum from patients with a clinically proven peanut allergy. Ara h 1 purified from peanuts heated at different temperatures exhibited IgE binding properties similar to those found for native Ara h 1, indicating that the allergenicity of Ara h 1 is heat-stable. We conclude that the allergenicity of Ara h 1 is unaffected by heating, although native Ara h 1 undergoes a significant heat-induced denaturation on a molecular level, indicating that the recognition of conformational epitopes of Ara h 1 by IgE either is not a dominant mechanism or is restricted to parts of the protein that are not sensitive to heat denaturation.
Koppelman
S J
SJ
TNO Nutrition and Food Research Institute, 3700 AJ Zeist, The Netherlands. koppelman@voeding.tno.nl
Bruijnzeel-Koomen
C A
CA
Hessing
M
M
de Jongh
H H
HH
D000328:Adult; D000485:Allergens / Q000737:chemistry*; D052179:Antigens, Plant; D010367:Arachis / Q000737:chemistry*; D002152:Calorimetry, Differential Scanning; D002850:Chromatography, Gel; D002942:Circular Dichroism; D006023:Glycoproteins; D006358:Hot Temperature; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood; D008565:Membrane Proteins; D010940:Plant Proteins / Q000737:chemistry*; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet