Department of Biological Science, Florida State University, Tallahassee, FL, USA.
Mol Immunol. 2013 Oct;55(3-4):253-63. doi: 10.1016/j.molimm.2013.02.004. Epub 2013 Mar 15.
Tree nuts are a widely consumed food. Although enjoyed safely by most individuals, allergic reactions to tree nuts, including almond, are not uncommon. Almond prunin (Pru du 6), an 11S globulin (legumin), is an abundant nut seed protein and a major allergen. Conformational epitope mapping studies of prunin have been performed with a murine monoclonal antibody (mAb) 4C10. This mAb reacts with non-reduced but not reduced prunin in immunoblotting assays, indicating the recognition of a conformational epitope. 4C10 competes with patient IgE, as assessed by ELISA, indicating clinical significance of the epitope. To characterize the 4C10 epitope, hydrogen/deuterium exchange (HDX) monitored by 14.5 T Fourier transform ion cyclotron resonance mass spectrometry (MS) was performed on the native prunin-4C10 complex and on uncomplexed native prunin. Several epitope candidate peptides that differ in deuterium uptake between the complexed and uncomplexed forms were identified. The epitope was further mapped by analyzing chimeric molecules incorporating segments of the homologous soybean allergen, Gly m 6, in immunoassays. These data indicate that the 4C10 epitope overlaps with a subset of patient IgE binding epitopes on almond prunin and further supports HDX-MS as a valid technique for mapping conformational epitopes.
坚果是一种广泛食用的食物。尽管大多数人都能安全享用，但对坚果的过敏反应并不少见，包括杏仁。杏仁朊（Pru du 6），一种 11S 球蛋白（legumin），是一种丰富的坚果种子蛋白，也是一种主要过敏原。已经使用鼠单克隆抗体（mAb）4C10 对朊进行了构象表位作图研究。该 mAb 在免疫印迹分析中与非还原但不还原的朊反应，表明识别构象表位。4C10 通过 ELISA 与患者 IgE 竞争，表明该表位具有临床意义。为了表征 4C10 表位，使用氢/氘交换（HDX）监测 14.5 T 傅里叶变换离子回旋共振质谱（MS）在天然朊-4C10 复合物和未结合的天然朊上进行。在复合物和未复合物之间，鉴定出几个在氘摄取方面存在差异的候选表位肽。通过在免疫分析中分析包含同源大豆过敏原 Gly m 6 片段的嵌合分子，进一步绘制了表位图。这些数据表明，4C10 表位与杏仁朊上一组患者 IgE 结合表位重叠，并进一步支持 HDX-MS 作为映射构象表位的有效技术。
