Molecular cloning, expression and characterization of Pru a 1, the major cherry allergen.
A high percentage of birch pollen allergic patients experiences food hypersensitivity reactions after ingestion of several fruits and vegetables. Previous work demonstrated common epitopes on an allergen of Mr 18,000 from sweet cherry (Prunus avium) and Bet v 1, the major allergen from birch pollen. N-terminal amino acid sequencing showed a sequence identity of 67% with Bet v 1. Here we report the cloning and cDNA sequencing of this cherry allergen. The entire deduced amino acid sequence described a protein of Mr 17,700 with 59.1% identity to Bet v 1. High degrees of identity in the range of 40 to 60% were also found with 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 into vector pET-16b and expressed in E. coli strain BL21(DE3) as a His-tag 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 allergosorbent 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 induction of a strong histamine release in basophils from cherry-allergic patients. Since sera from 17/19 of such patients contained IgE against this 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 extract and hence could be a useful tool for studying the molecular and immunological properties of pollen related food allergens.
Scheurer
S
S
Paul-Ehrlich-Institute, Langen, Germany.
Metzner
K
K
Haustein
D
D
Vieths
S
S
D000485:Allergens / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D001483:Base Sequence; D003001:Cloning, Molecular; D005638:Fruit / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D015870:Gene Expression; D017343:Genes, Plant; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D016415:Sequence Alignment
D000485:Allergens; D052179:Antigens, Plant; C109472:PRUA1 protein, Prunus avium; D010940:Plant Proteins; C061312:Bet v 1 allergen, Betula; D007073:Immunoglobulin E