Comparative studies on tree-pollen allergens. VI. The effect of thermal denaturation on the reactivity of birch pollen (Betula verrucosa) allergens.

The structural changes as due to thermal denaturation of antigens and allergens of birch pollen produced by gel filtration chromatography were studied by circular dichroism, RAST inhibition and immunoelectrophoretic techniques. Results of circular dichroism showed that both BV3 and BV4 contained 20-25% alfa-helix, while BV45 consists of beta-pleated sheet and random coil. Fractions BV3 and BV4 lost about 50% of their native tertiary structures when heated at 100 degrees C for 3 h. The number of antigenic lines found in the CIE plates for the heat-treated fractions was reduced as well. The CRIE preparations showed unchanged radiostaining for the heated BV3 fraction (correlated to the untreated BV3 fraction). For the BV4 fraction a diminished radiostaining was observed. A reduction of 8-fold and 4-fold of the IgE binding capacity was observed in RAST inhibition for fractions BV3 and BV4, respectively after heating. The purified fraction BV45 showed 50% decrease in CD absorption after heating to 100 degrees C for 1 h. The CIE pattern of the heated fraction gave a single precipitate line and the area under the precipitate line was reduced as compared to the unheated fraction. The CRIE plate showed reduced intensity of radiostaining. Skin prick tests and PK tests confirmed the reduced allergenicity of the heated fractions. These findings confirm the relative thermostability of the allergens in birch pollen. However, heating infers irreversible changes on the tertiary structure of the molecules, and these structural changes can be correlated to a quantitative reduction of allergenic reactivity.