Akagawa Mitsugu, Handoyo Tri, Ishii Takeshi, Kumazawa Shigenori, Morita Naofumi, Suyama Kyozo
Department of Biological Chemistry, Division of Applied Life Science, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Sakai 599-8531, Japan.
J Agric Food Chem. 2007 Aug 22;55(17):6863-70. doi: 10.1021/jf070843a. Epub 2007 Jul 27.
Wheat can cause severe IgE-mediated systematic reactions, but knowledge on relevant wheat allergens at the molecular level is scanty. The aim of the present study was to achieve a more detailed and comprehensive characterization of the wheat allergens involved in food allergy to wheat using proteomic strategies, referred to as "allergenomics". Whole flour proteins were separated by two-dimensional gel electrophoresis with isoelectric focusing and lithium dodecyl sulfate-polyacrylamide gel electrophoresis. Then, IgE-binding proteins were detected by immunoblotting with sera of patients with a food allergy to wheat. After tryptic digestion, the peptides of IgE-binding proteins were analyzed by matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry. In this study, we identified four previously reported wheat allergens or their sequentially homologous proteins [serpin, alpha-amylase inhibitor, gamma-gliadin, and low molecular weight (LMW) glutenin] by a database search. As a result of the high resolution of two-dimensional gel electrophoresis, nine subunits of LMW glutenins were identified as the most predominant IgE-binding antigens. The two-dimensional allergen map can be beneficial in many ways. It could be used, for example, for precise diagnosis of wheat-allergic patients and assessment of wheat allergens in food. Additionally, we compared allergenomics to conventional biochemical methods and evaluated the usefulness of a proteomic strategy for identifying putative allergens to wheat allergy.
小麦可引发严重的免疫球蛋白E（IgE）介导的全身反应，但在分子水平上有关小麦相关过敏原的知识却很匮乏。本研究的目的是利用蛋白质组学策略（即“过敏原组学”），更详细、全面地表征与小麦食物过敏相关的小麦过敏原。通过等电聚焦和十二烷基硫酸锂-聚丙烯酰胺凝胶电泳的二维凝胶电泳对全麦粉蛋白质进行分离。然后，用对小麦食物过敏患者的血清进行免疫印迹检测IgE结合蛋白。胰蛋白酶消化后，通过基质辅助激光解吸电离串联飞行时间质谱分析IgE结合蛋白的肽段。在本研究中，我们通过数据库搜索鉴定出四种先前报道的小麦过敏原或其序列同源蛋白[丝氨酸蛋白酶抑制剂、α-淀粉酶抑制剂、γ-醇溶蛋白和低分子量（LMW）谷蛋白]。由于二维凝胶电泳的高分辨率，LMW谷蛋白的九个亚基被鉴定为最主要的IgE结合抗原。二维过敏原图谱在许多方面都有益处。例如，它可用于小麦过敏患者的精确诊断以及食品中小麦过敏原的评估。此外，我们将过敏原组学与传统生化方法进行了比较，并评估了蛋白质组学策略在鉴定小麦过敏潜在过敏原方面的实用性。
