CLONING AND SEQUENCING OF A GENE ENCODING A 2S ALBUMIN SEED STORAGE PROTEIN-PRECURSOR FROM ENGLISH WALNUT (JUGLANS-REGIA), A MAJOR FOOD ALLERGEN

Background: Walnuts rank third in per capita consumption of tree nuts in the United States and can be associated with systemic IgE-mediated reactions in some individuals. Objective: The objectives of the study were to clone a gene encoding one of the major food allergens in the w alnut kernel and to characterize the recombinant allergen, Methods: A cDNA expression library in the lambda vector Uni-ZAP, which was prepar ed from walnut somatic embryos, was screened by using a patient's sera that reacted with multiple protein bands on immunoblotting. Results: A cDNA clone containing an insert of 663 bp was identified and named J ug r I. DNA sequence analysis of this clone revealed that it encoded a protein 142 amino acids in length, Comparison of the encoded protein sequences with protein databases revealed that this clone exhibits a 4 6.1% identity with the Brazil nut (Bertholletia excelsa) methionine-ri ch 2S albumin seed storage protein precursor, Ber e 1. Jug r 1 appears to be an important walnut food allergen; 12 of 16 sera from patients allergic to walnuts demonstrated IgE binding to the 2S albumin seed st orage protein precursor fusion protein. An IgE-binding inhibition stud y suggests that the walnut 28 protein precursor undergoes posttranslat ional modification into a large and small subunit that is similar to c astor seed, cottonseed, mustard seed, and Brazil nut 28 seed storage p rotein allergens, Interestingly, the gene encoding this allergenic pro tein in Brazil nuts has recently gained notoriety because of its exper imental use as a transgene to enhance the nutritional quality of legum es. Conclusion: This is now the sixth definitive 28 albumin seed stora ge protein demonstrated to bind IgE, suggesting that this class of pro teins is inherently allergenic.