Hales Belinda J, Bosco Anthony, Mills Kristina L, Hazell Lee A, Loh Richard, Holt Patrick G, Thomas Wayne R
Centre for Child Health Research, University of Western Australia, Telethon Institute for Child Health Research, West Perth, Australia.
Int Arch Allergy Immunol. 2004 Oct;135(2):101-7. doi: 10.1159/000080652. Epub 2004 Sep 2.
The major peanut allergen Ara h 2 consists of two isoforms, namely Ara h 2.0101 and Ara h 2.0201. The recently identified Ara h 2.0201 isoform contains an extra 12 amino acids including an extra copy of the reported immunodominant epitope DPYSPS. This study aimed to evaluate the IgE binding of the two Ara h 2 isoforms.
Ten clones of Ara h 2 were sequenced to assess the relative frequency of the Ara h 2 isoforms and to identify whether there was further variation in the Ara h 2 sequence. IgE binding to Ara h 2.0101 and Ara h 2.0201 was measured for 70 peanut-allergic children using an IgE DELFIA assay to quantitate specific IgE binding. A competition assay was used to measure whether Ara h 2.0201 contained IgE epitopes other than those found for Ara h 2.0101.
The original Ara h 2.0101 sequence was found for 6/10 clones and Ara h 2.0201 was found for 2/10 clones. Ara h 2.0201 had the expected insertion of 12 amino acids as well as substitutions at positions 40 (40G) and 142 (142E). Two new isoforms were identified as different polymorphisms of position 142. One Ara h 2.01 clone (Ara h 2.0102) contained 142E and one Ara h 2.02 clone (Ara h 2.0202) contained 142D. A polymorphism that was previously identified by other investigators at position 77 (77Q or 77R) was not found for any of the 10 sequences. Although the level of IgE binding to Ara h 2.0201 of individual patients was frequently higher than the binding to Ara h 2.0101 (p < 0.01), there was a strong correlation in binding to both isoforms (r = 0.987, p < 0.0001) and when analyzed as a group the means were similar. Ara h 2.0101 was not as efficient at blocking reactivity to Ara h 2.0201 indicating there is an additional IgE specificity for the Ara h 2.0201 isoform.
Ara h 2.0201 has similar but higher IgE binding than the originally sequenced Ara h 2.0101 isoform and contains other IgE specificities.
主要花生过敏原Ara h 2由两种亚型组成，即Ara h 2.0101和Ara h 2.0201。最近鉴定出的Ara h 2.0201亚型包含额外的12个氨基酸，其中包括已报道的免疫显性表位DPYSPS的一个额外拷贝。本研究旨在评估这两种Ara h 2亚型的IgE结合情况。
对10个Ara h 2克隆进行测序，以评估Ara h 2亚型的相对频率，并确定Ara h 2序列中是否存在进一步的变异。使用IgE DELFIA检测法对70名花生过敏儿童测定其与Ara h 2.0101和Ara h 2.0201的IgE结合，以定量特异性IgE结合。采用竞争检测法测量Ara h 2.0201是否含有除Ara h 2.0101中发现的IgE表位以外的其他IgE表位。
在10个克隆中，6个克隆发现了原始的Ara h 2.0101序列，2个克隆发现了Ara h 2.0201。Ara h 2.0201具有预期的12个氨基酸插入以及40位（40G）和142位（142E）的替换。鉴定出两种新亚型为142位的不同多态性。一个Ara h 2.01克隆（Ara h 2.0102）含有142E，一个Ara h 2.02克隆（Ara h 2.0202）含有142D。在10个序列中均未发现先前其他研究者在77位（77Q或77R）鉴定出的多态性。尽管个体患者与Ara h 2.0201的IgE结合水平通常高于与Ara h 2.0101的结合水平（p < 0.01），但与两种亚型的结合存在强相关性（r = 0.987，p < 0.0001），并且作为一个整体分析时，平均值相似。Ara h 2.0101在阻断对Ara h 2.0201的反应性方面效率不高，表明Ara h 2.0201亚型存在额外的IgE特异性。
Ara h 2.0201与最初测序的Ara h 2.0101亚型具有相似但更高的IgE结合，并且含有其他IgE特异性。
