Pomegranate (Punica granatum L.) Expresses Several nsLTP Isoforms Characterized by Different Immunoglobulin E-Binding Properties
Pomegranate (Punica granatum L.) Expresses Several nsLTP Isoforms Characterized by Different Immunoglobulin E-Binding Properties
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DOI：
10.1159/000362761
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发表时间：
2014-01-01
影响因子：
2.8
通讯作者：
Zoccatelli, Gianni
中科院分区：
文献类型：
--
作者：
Bolla, Michela;Zenoni, Sara;Zoccatelli, Gianni
背景：石榴过敏与对非特异性脂转移蛋白(NsLTPs)的致敏有关。我们的目的是在分子水平上鉴定和鉴定石榴中表达的非特异性nsLTP，并从免疫球蛋白E(IgE)结合和与桃nsLTP(PrU P3)的交叉反应方面研究它们的致敏特性。方法：建立了酸尿素PAGE和十二烷基硫酸钠PAGE非平衡双向电泳法分离石榴nsLTP。用抗PrUP3多克隆抗体和石榴过敏患者血清进行免疫印迹试验，检测其免疫反应性。石榴nsLTPs经层析纯化后，经胰酶消化和质谱仪(MS)分析进行最终鉴定。为此，通过克隆三个石榴nsLTP的cDNA获得的序列被整合到数据库中，随后搜索用于MS数据解释。结果：2-D免疫印迹法鉴定出4个nsLTP。克隆和MS分析表明，3种NsLTP异构体的氨基酸序列同源性为66-68%，分别为PUN g 1.0101、PUN g 1.0201和PUN g 1.0301。结论：通过双向电泳法可以分离出具有不同IgE结合特性的不同nsLTP亚型，这可能反映了特异性的致敏性。Pru p3对起始敏化的贡献并不像其他植物nsLTP那样是中心的。(C)2014年S.Karger AG，巴塞尔
Background: Pomegranate allergy is associated with sensitization to non-specific lipid transfer proteins (nsLTPs). Our aim was to identify and characterize the non-specific nsLTPs expressed in pomegranate at the molecular level and to study their allergenic properties in terms of immunoglobulin E (IgE)-binding and cross-reactivity with peach nsLTP (Pru p 3). Methods: A non-equilibrium two-dimensional (2-D) electrophoretic approach based on acid-urea PAGE and sodium dodecyl sulfate PAGE was set up to separate pomegranate nsLTPs. Their immunoreactivity was tested by immunoblotting carried out with anti-Pru p 3 polyclonal antibodies and sera from pomegranate-allergic patients. For final identification, pomegranate nsLTPs were purified by chromatography and subjected to trypsin digestion and mass spectrometry (MS) analysis. For this purpose, the sequences obtained by cDNA cloning of three pomegranate nsLTPs were integrated in the database that was subsequently searched for MS data interpretation. Results: Four nsLTPs were identified by 2-D immunoblotting. The detected proteins showed different IgE-binding capacity and partial cross-reactivity with Pru p 3. cDNA cloning and MS analyses led to the identification of three nsLTP isoforms with 66-68% amino acid sequence identity named Pun g 1.0101, Pun g 1.0201 and Pun g 1.0301. Conclusions: By 2-D electrophoresis, we could separate different nsLTP isoforms possessing different IgE-binding properties, which might reflect peculiar allergenic potencies. The contribution of Pru p 3 to prime sensitization is not central as in other plant nsLTPs. (C) 2014 S. Karger AG, Basel