Bd 30K Research Articles

Reduction of the allergenicity of soybean by treatment with proteases.

Soybeans were treated with proteases to reduce allergenicity. By immunoblotting with a monoclonal antibody against a major soybean allergen (Gly m Bd 30K), two of eight proteases so far tested were selected to achieve a reduction in allergenicity. Both antigenicity to the monoclonal antibody and allergenicity to the sera from soybean-sensitive patients proved to be markedly reduced by processing with either protease. Thus, soybeans treated with an appropriate protease may possibly be supplied as a hypoallergenic foodstuff for patients.

Epitope analysis of soybean major allergen Gly m Bd 30K recognized by the mouse monoclonal antibody using overlapping peptides.

The epitopes of the major soybean allergen, Gly m Bd 30K, recognized by mouse monoclonal antibodies H6 and F5 were investigated by using synthetic peptides bound to pins. The epitopes are shown to be localized in peptide 31QGGCGRGWAFSATGAIEA48 for H6, and in 115DKVTIDGYETLIMSDEST132 for F5.

Removal of mite allergens from blankets: Comparison of dry cleaning and hot water washing

1. Barry G, Kishore G, Padgette S, et al. Inhibitors of amino acid biosynthesis: strategies for imparting glyphosate tolerance to crop plants. In: Singh BK, Flores HE, Shannon JC, eds. Biosynthesis and molecular regulation of amino acids in plants. Rockville, Maryland: American Society of Plant Physiologists, 1992:139-45. 2. Padgette SR, Re DB, Barry GF, et al. New weed control opportunities: development of soybeans with a Roundup Ready TM gene. In: Duke SO, ed. Herbicide-resistant crops: agricultural economic, environmental, regulatory, and technological aspects. CRC Press, 1994 (in press). 3. Burks AW, Brooks JR, Sampson HA. Allergenicity of major component proteins of soybean determined by enzyme linked immunosorbent assay (ELISA) and immunoblotting in children with atopic dermatitis and positive soy challenges. J ALLERGY CLIN IMMUNOL 1988;81:1135-42. 4. Thanh VH, Shibasaki K. Major proteins of soybean seeds: a straight forward fractionation and their characterization. J Agri Food Chem 1976;24:1117-21. 5. Ogawa F, Tsuji H, Bando N, et al. Identification of the soybean allergenic protein, Gly m Bd 30K, with the soybean seed 34-kDa oil-body associated protein. Biosci Biotech Biochem 1993;57:1030-3.

Measurement ofGly mBd 30K, a Major Soybean Allergen, in Soybean Products by a Sandwich Enzyme-linked Immunosorbent Assay

By a sandwich enzyme-linked immunosorbent assay, a soybean major allergen, Gly m Bd 30K, in soybean products was measured. The allergen occurred at high concentrations in soy milk, tofu, kori-dofu, and yuba, but its content in kinako was small. No allergen was found in fermented foods such as miso, shoyu, and natto. The allergen was clearly shown to occur in meat balls, beef croquettes, and fried chicken that contained soybean protein isolate.

Reduction of the Soybean Allergenicity by the Fermentation with Bacillus natto.

We investigated the effects of the fermentation with Bacillus natto on the allergenicity ascribed to the major soybean allergen, Gly m Bd 30K. Time dependent decreases in 1) reactivity for IgE antibody from a soybean-sensitive patient and 2) reactivity for a monoclonal antibody were observed during fermentation, and the electrophoretic patterns of the proteins in the fermented soybean showed all the proteins were hydrolyzed to the small fragments with molecular weight below 10,000. These results suggested that the epitopes of the Gly m Bd 30K against the mouse monoclonal IgG and human serum IgE were decomposed by the digestion with the proteases of Bacillus natto during fermentation and that natto might be a hypoallergenic soybean product.

Identification of the soybean allergenic protein, Gly m Bd 30K, with the soybean seed 34-kDa oil-body-associated protein.

The soybean allergenic protein, Gly m Bd 30K [Ogawa et al., J. Nutr. Sci. Vitaminol., 37, 555-565 (1991)] which is most strongly and frequently recognized by the IgE antibodies in sera of soybean-sensitive patients with atopic dermatitis, has been characterized. The allergen was isolated from the crude 7S-globulin fraction as an oligomeric form with a molecular weight of more than 3000,000 by gel-filtration chromatography. On two-dimensional gel electrophoresis, the native oligomeric allergen had an isoelectric point of about pH 4.5 and was dissociated into a monomeric form with a molecular weight of about 32,000 by the treatment with sodium dodecyl sulfate and 2-mercaptoethanol. The monomeric allergen had an N-terminal amino acid sequence and amino acid composition identical with those of the soybean seed 34-kDa oil-body-associated protein or the soybean vacuolar protein P34 with close homology to papain-like thiol proteinases [Kalinski et al., J. Biol. Chem., 267, 12068 (1992)]. The identity was further confirmed by the immunological cross-reactivity to the antibodies produced against each of the purified allargen and the 34-kDa oil-body-associated protein. By this observation, Gly m Bd 30K was shown to have about 30% sequence homology with Der pI, a house dust mite allergen that is a thiol proteinase from Dermatophagoides pteronyssius.

Preparation and application of monoclonal antibodies for a sandwich enzyme-linked immunosorbent assay of the major soybean allergen, Gly m Bd 30K.

In order to obtain probes suitable for the determination of the major soybean allergen, Gly m Bd 30 K, in soybean-related processed foods, we have prepared two monoclonal antibodies, F5 of IgG2a and H6 of IgM, by the fusion of P3U1 myeloma cells with the spleen cells of BALB/c mice immunized with the reductively carboxymethylated allergen (RCM-allergen). The two monoclonal antibodies were shown to be specific to the intact allergen as well as the RCM-allergen and to recognize distinct epitopes on the allergen. Of the monoclonal antibodies, F5 was conjugated with peroxidase. H6 and the labeled F5 were applied as the fixing antibody and the labeled first antibody, respectively, for a direct sandwich enzyme-linked immunosorbent assay of the allergen. When the allergen was extracted from the soybean-related foods with Tris-HCl buffer containing sodium dodecylsulfate and mercaptoethanol, the allergen, Gly m Bd 30 K, was shown to be measured in a range of 5-500 ng in this assay.