Cellular and molecular characterization of a major soybean allergen.
Soybean proteins share a large number of cross-reacting allergens with other members of the legume family; however, soy-allergic patients rarely react clinically to other members of the legume family. Gly m Bd 30K, an IgE-binding protein with a molecular weight of 30 kD, was identified in soybean extracts by Western IgE-immunoblot analysis. This monomeric allergen was shown to have an N-terminal amino acid sequence and amino acid composition identical to that of the seed 34-kD protein, P34, a thiol protease of the papain family. Electron-microscopic immunolocalization of P34 monoclonal antibodies and IgE binding to sections of soybean seeds showed dense staining throughout the vacuolar bodies, localizing the allergens in protein storage vacuoles of seed cotyledons. We used pooled serum from soybean-sensitive patients to determine the linear IgE-specific epitopes in the 34-kD allergen amino acid sequence. B-cell epitope mapping revealed 10 regions of IgE-binding activity using an overlapping peptide strategy of 15-mers offset by 8 amino acids throughout the P34 sequence. Smaller overlapping peptides, 10-mers offset by 2 amino acids, revealed 16 distinct linear epitopes, 9 of which were mapped to the mature protein. No obvious amino acid sequence motifs could be identified by the smaller IgE-binding epitopes. Using individual patient serum, 5 immunodominant epitopes were identified in this allergen.
Helm
R
R
Department of Pediatrics, University of Arkansas for Medical Sciences, Arkansas Children's Hospital Research Institute, Little Rock, Ark., USA.
Cockrell
G
G
Herman
E
E
Burks
A
A
Sampson
H
H
Bannon
G
G
D000485:Allergens / Q000737:chemistry*; D000595:Amino Acid Sequence; D039741:Antigens, Dermatophagoides; D052179:Antigens, Plant; D018548:Cotyledon / Q000737:chemistry / Q000648:ultrastructure; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000276:immunology; D018604:Epitope Mapping; D006023:Glycoproteins / Q000737:chemistry / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D030262:Soybean Proteins; D013025:Glycine max / Q000737:chemistry / Q000276:immunology*; D014617:Vacuoles / Q000648:ultrastructure
D000485:Allergens; D039741:Antigens, Dermatophagoides; D052179:Antigens, Plant; C090803:Gly m I protein, Glycine max; D006023:Glycoproteins; D010940:Plant Proteins; D030262:Soybean Proteins; C075135:P34 protein, Glycine max; D007073:Immunoglobulin E; D003546:Cysteine Endopeptidases