Hiemori Miki, Ito Hitomi, Kimoto Masumi, Yamashita Hiromi, Nishizawa Keito, Maruyama Nobuyuki, Utsumi Shigeru, Tsuji Hideaki
Department of Nutritional Science, Faculty of Health and Welfare Science, Okayama Prefectural University, Kuboki 111, Soja 719-1197, Japan.
Biochim Biophys Acta. 2004 Nov 18;1675(1-3):174-83. doi: 10.1016/j.bbagen.2004.09.003.
One of the major soybean allergens, Gly m Bd 28K, is suggested to be biosynthesized as a preproprotein form, which would be composed of a signal peptide, Gly m Bd 28K and the C-terminal peptide (the 23-kDa peptide). However, the 23-kDa peptide has never been characterized. In the present study, we prepared a monoclonal antibody (mAb) against a recombinant 23-kDa peptide expressed in Escherichia coli to detect the 23-kDa peptide in soybean. Several proteins were detected by immunoblotting with the mAb. All of the proteins were shown to have the identical N-terminal amino acid sequence, suggesting that the proteins correspond to the C-terminal part of the Gly m Bd 28K precursor. Furthermore, Gly m Bd 28K and the 23-kDa peptide were observed to come out at the 21st day after flowering and to locate in the crystalloid part of protein storage vacuoles in growing cotyledons. Some of the 23-kDa peptides were shown to be glycoproteins with an N-linked glycan moiety and exhibited the binding to IgE antibodies in the sera of patients sensitive to soybean. The binding of the peptides to IgE antibodies was suggested to be predominantly dependent on their glycan moiety. This study proves the occurrence of the 23-kDa peptide in soybean and that it is a new allergen.
大豆主要过敏原之一Gly m Bd 28K被认为是以前原蛋白形式生物合成的，它由一个信号肽、Gly m Bd 28K和C末端肽（23 kDa肽）组成。然而，23 kDa肽从未被鉴定过。在本研究中，我们制备了一种针对在大肠杆菌中表达的重组23 kDa肽的单克隆抗体（mAb），以检测大豆中的23 kDa肽。用该单克隆抗体通过免疫印迹检测到了几种蛋白质。所有这些蛋白质都显示出相同的N末端氨基酸序列，这表明这些蛋白质对应于Gly m Bd 28K前体的C末端部分。此外，观察到Gly m Bd 28K和23 kDa肽在开花后第21天出现，并定位于正在生长的子叶中蛋白质储存液泡的晶体部分。一些23 kDa肽被证明是具有N-连接聚糖部分的糖蛋白，并在对大豆敏感的患者血清中表现出与IgE抗体的结合。肽与IgE抗体的结合主要被认为取决于它们的聚糖部分。本研究证实了大豆中存在23 kDa肽，并且它是一种新的过敏原。
