Significance of 40-, 45-, and 48-kDa Proteins in the Moderate-to-Severe Clinical Symptoms of Buckwheat Allergy

PURPOSE: This study was aimed to investigate the relationship between the allergen components and moderate-to-severe allergic reactions in patients with buckwheat allergy. METHODS: Fifteen patients with a history of buckwheat ingestion and a buckwheat specific IgE level> or =0.35 kU/L were enrolled. They were divided into 2 groups according to clinical severity scores, with 0-1 being asymptomatic-to-mild and 2-4 being moderate-to-severe symptoms. Immunoblotting was performed to investigate IgE reactivity toward buckwheat allergens and to measure intensity of each component by using a reflective densitometer. RESULTS: The proportions of positive band to the 16 kDa (62.5% vs 0%, P=0.026) and 40-50 kDa (87.5% vs 28.6%, P=0.041) buckwheat allergens in the grade 2-4 group were higher than those in grade 0-1 group. The level of buckwheat specific IgE of grade 2-4 group was higher than that of grade 0-1 group (41.3 kU/L vs 5.5 kU/L, P=0.037). The median optical densities (ODs) of IgE antibody binding to 40-50 kDa protein were higher in the grade 2-4 group, compared with those in the grade 0-1 group (130% OD vs 60.8% OD, P=0.037). CONCLUSIONS: The 40-50 kDa protein is implicated as an important allergen to predict moderate-to-severe clinical symptoms in Korean children with buckwheat allergy.

Erratum: Effects of enzymatic hydrolysis of buckwheat protein on antigenicity and allergenicity

We made a mistake in presenting author affiliations.

Effects of enzymatic hydrolysis of buckwheat protein on antigenicity and allergenicity

BACKGROUND/OBJECTIVES: Due to its beneficial health effects, use of buckwheat has shown a continuous increase, and concerns regarding the allergic property of buckwheat have also increased. This study was conducted for evaluation of the hydrolytic effects of seven commercial proteases on buckwheat allergens and its allergenicity. MATERIALS/METHODS: Extracted buckwheat protein was hydrolyzed by seven proteolytic enzymes at individual optimum temperature and pH for four hours. Analysis was then performed using SDS-PAGE, immunoblotting, and competitive inhibition ELISA (ciELISA) with rabbit antiserum to buckwheat protein, and direct ELISA with pooled serum of 21 buckwheat-sensitive patients. RESULTS: Alkaline protease, classified as serine peptidase, was most effective in reducing allergenicity of buckwheat protein. It caused decomposition of the whole buckwheat protein, as shown on SDS-PAGE, and results of immunoblotting showed that the rabbit antiserum to buckwheat protein no longer recognized it as an antigen. Allergenicity showed a decrease of more than 50% when pooled serum of patients was used in ELISA. Two proteolytic enzymes from Aspergillus sp. could not hydrolyze buckwheat allergens effectively, and the allergenicity even appeared to increase. CONCLUSIONS: Serine-type peptidases appeared to show a relatively effective reduction of buckwheat allergenicity. However, the antigenicity measured using rabbit antiserum did not correspond to the allergenicity measured using sera from human patients. Production of less allergenic buckwheat protein may be possible using enzymatic hydrolysis.

Enhancement of Protein Productivity of Recombinant Hepatitis A Virus VP1 in Stably Transfected Drosophila melanogaster S2 Cells

The effect of DMSO and sodium butyrate on the production of recombinant hepatitis A virus (HAV) capsid protein VP1 was evaluated and optimized in the culture of stably transfected Drosophila melanogaster S2 cells using culture plates and spinner flasks. The effect of DMSO and sodium butyrate was also evaluated to improve the recombinant VP1 production in stably transfected Drosophila S2 cells. A production level of 0.88 mg of recombinant VP1/liter was obtained in the culture-plate culture of stably transfected S2 cells at 6 days after induction with 0.5 mM CuSO4. The supplements of 2% DMSO and 10 mM sodium butyrate at 4 days post-inoculation increased recombinant VP1 accumulation by 141 and 104%, respectively, resulting in 2.17 and 1.7 mg/liter of recombinant VP1 production. In spinner flasks, recombinant VP1 production reached maximum level at 9 days after induction with 0.5 mM CuSO4, with approximately 4.96 mg/liter of recombinant VP1 production level. When 2% DMSO or 10 mM sodium butyrate was added at 5 days post-inoculation, the recombinant VP1 production was increased to 8.35 and 5.85 mg/liter, respectively. However, the synergistic effects of DMSO and sodium butyrate were not observed. These results indicate that DMSO and/or sodium butyrate can be successfully used to improve the recombinant HAV VP1 production in culture plates and spinner flasks.