New strategies for allergen-specific immunotherapy have focused on reducing IgE reactivity of purified recombinant allergens while maintaining T-cell epitopes. Previously, we showed that disrupting the disulfide bonds of the major house dust mite allergen Der p 2 resulted in 10-100-fold less skin test reactivity in mite-allergic subjects but did not change in vitro T-cell proliferative responses. To provide a more complete picture of the antigenic surface of Der p 2, we report here the identification of three epitopes using hydrogen protection nuclear magnetic resonance spectroscopy. The epitopes are defined by monoclonal antibodies that are able to inhibit IgE antibody binding to the allergen. Each monoclonal antibody affected the amide exchange rate of 2-3 continuous residues in different regions of Der p 2. Based on these data, a number of other residues were predicted to belong to each epitope, and this prediction was tested for monoclonal antibody 7A1 by generating alanine point mutants. The results indicate that only a small number of residues within the predicted epitope are functionally important for antibody binding. The molecular definition of these three epitopes will enable us to target limited positions for mutagenesis and to expand our studies of hypoallergenic variants for immunotherapy.

中文翻译：

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氢交换核磁共振光谱图绘制了屋尘螨过敏原Der p 2上的抗体表位。

变应原特异性免疫疗法的新策略集中于降低纯化的重组变应原的IgE反应性，同时保持T细胞表位。以前，我们表明破坏主要的室内尘螨过敏原Der p 2的二硫键可导致螨过敏受试者的皮肤测试反应性降低10-100倍，但不会改变体外T细胞增殖反应。为了提供Der p 2抗原表面的更完整图片，我们在此报告使用氢保护核磁共振波谱鉴定三个表位。表位由能够抑制IgE抗体与变应原结合的单克隆抗体定义。每种单克隆抗体都会影响Der p 2不同区域中2-3个连续残基的酰胺交换速率。基于这些数据，预测许多其他残基属于每个表位，并通过生成丙氨酸点突变体对该单克隆抗体7A1进行了测试。结果表明，预测表位中只有少量残基在功能上对抗体结合很重要。这三个表位的分子定义将使我们能够针对诱变的有限位置进行定位，并扩大我们对用于免疫疗法的低变应原变体的研究。