MOLECULAR-CLONING, EXPRESSION AND CHARACTERIZATION OF PRU A-1, THE MAJOR CHERRY ALLERGEN

A high percentage of birch pollen allergic patients experiences food h ypersensitivity reactions after ingestion of several fruits and vegeta bles. Previous work demonstrated common epitopes on an allergen of M-r 18 000 from sweet cherry (Prunus avium) and Bet v 1, the major allerg en from birch pollen. N-terminal amino acid sequencing showed a sequen ce identity of 67% with Bet v 1. Here we report the cloning and cDNA s equencing of this cherry allergen. The entire deduced amino acid seque nce described a protein of M-r 17 700 with 59.1% identity to Bet v 1. High degrees of identity in the range of 40 to 60% were also found wit h related allergens from other kinds of tree pollen and plant foods as well as with stress-induced proteins from food plants such as parsley , potato and soya. The coding DNA of the cherry protein was cloned int o vector pET-1Gb and expressed in E. coli strain BL21(DE3) as a His-ta g fusion protein. As shown by SDS-PAGE, the apparent molecular masses of the nonfusion protein and the natural allergen were identical. The fusion protein showed high IgE binding potency when sera from patients allergic to cherry were tested by immunoblotting and enzyme allergoso rbent tests. Moreover, it cross-reacted strongly with IgE specific for the natural counterpart and for Bet v 1. The high biological activity of the recombinant fusion protein was further confirmed by the induct ion of a strong histamine release in basophils from cherry-allergic pa tients. Since sera from 17/19 of such patients contained IgE against t his allergen it was classified as a major allergen and named Pru a 1. Recombinant Pru a 1 mimics most of the allergenic potency of cherry ex tract and hence could be a useful tool for studying the molecular and immunological properties of pollen related food allergens. (C) 1997 Pu blished by Elsevier Science Ltd.