Development of an inexpensive, simple purification method for ω-5 gliadin from wheat flour

IF 2.9 Q2 MULTIDISCIPLINARY SCIENCES

MethodsX Pub Date : 2026-06-01 DOI:10.1016/j.mex.2026.103979

Akihiro Maeta , Mamoru Tanaka , Kyoko Takahashi

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Abstract

ω-5 gliadin is a major causative allergen associated with various types of wheat allergy, including wheat-dependent exercise-induced anaphylaxis. Recently, ω-5 gliadin-deficient wheat has been developed. However, several wheat protein studies do not use standard ω-5 gliadin because its naturally derived form is expensive, and purifying it in-house is time-consuming. Therefore, we sought to develop an inexpensive, simple method for purifying ω-5 gliadin on a small scale.

▪
We compared ω-5 gliadin-deficient wheat with normal wheat and screened for ω-5 gliadin bands using acid-polyacrylamide gel electrophoresis (A-PAGE) on a mini-slab electrophoresis apparatus.
▪
The ω-5 gliadin band was excised and sealed in dialysis tubing, and the protein was electroeluted using agarose gel electrophoresis. After electroelution, the eluted solution was filtered, dialyzed, and then freeze-dried.
▪
The main band of collected proteins migrated at approximately 55 kDa in SDS-PAGE, and the most intense band was identified as ω-5 gliadin using nano-LC-MS/MS.

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一种廉价、简单的从小麦粉中提纯ω-5麦胶蛋白的方法

ω-5麦胶蛋白是与多种小麦过敏相关的主要致敏原，包括小麦依赖性运动致过敏反应。近年来，ω-5麦胶蛋白缺乏小麦已被开发出来。然而，一些小麦蛋白研究没有使用标准的ω-5麦胶蛋白，因为它的自然衍生形式是昂贵的，并且在内部纯化它是耗时的。因此，我们寻求一种廉价、简单的方法来小范围提纯ω-5麦胶蛋白。▪我们将ω-5麦胶蛋白缺乏的小麦与正常小麦进行比较，并在微型平板电泳仪上使用酸-聚丙烯酰胺凝胶电泳（a - page）筛选ω-5麦胶蛋白带。▪切除ω-5麦胶蛋白带，在透析管中密封，琼脂糖凝胶电泳电泳蛋白。电洗脱后，将洗脱液过滤，透析，然后冷冻干燥。▪收集到的蛋白主带在SDS-PAGE中迁移约55 kDa，并通过纳米lc -MS/MS鉴定为ω-5麦胶蛋白。

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