Barderas Rodrigo, Purohit Ashok, Rodríguez Rosalía, Pauli Gabrielle, Villalba Mayte
Departmento Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, Spain.
Ann Allergy Asthma Immunol. 2006 Apr;96(4):557-63. doi: 10.1016/S1081-1206(10)63550-8.
Fra e 1 is a major allergen for ash pollen-sensitized individuals in northern and central Europe. It belongs to the Ole e 1-like family and displays high cross-reactivity with taxonomically related members.
To isolate and characterize natural Fra e 1 (nFra e 1) from ash pollen and to compare its structural, antigenic, and allergenic properties with those of its recombinant form (rFra e 1).
The allergen was isolated by means of gel permeation chromatography and reverse-phase high-performance liquid chromatography columns. Molecular characterization was performed by means of Edman degradation, mass spectrometry, circular dichroism, concanavalin A lectin reaction, and anti-horseradish peroxidase polyclonal antibody. Immunologic characterization was performed using immunoblotting and enzyme-linked immunosorbent assay, inhibition experiments, and histamine release assays with serum samples from allergic patients with well-known reactivity to Fra e 1 or Ole e 1 and with polyclonal antiserum and monoclonal antibodies against Ole e 1. The protein used as a reference was rFra e 1, which was produced in the yeast Pichia pastoris.
Purified nFra e 1 appeared as 5 variants with different glycosylation degrees. Both nFra e 1 and rFra e 1 were equivalently folded as deduced from the spectroscopic analysis using circular dichroism. Both molecules share the antigenic and allergenic epitopes after the purification process, and the glycan group of nFra e 1 is a potential epitope. Natural Fra e 1 displayed strong cross-reactivity with Ole e 1.
Natural Fra e 1 is a heterogeneously glycosylated protein with high allergenic relevance. It displays structural, antigenic, and allergenic similarity with rFra e 1. Both proteins could be used for clinical purposes.
Fra e 1是北欧和中欧对白蜡花粉过敏个体的主要过敏原。它属于Ole e 1样家族，与分类学上相关的成员表现出高度交叉反应性。
从白蜡花粉中分离并鉴定天然Fra e 1（nFra e 1），并将其结构、抗原性和过敏原性与重组形式（rFra e 1）进行比较。
通过凝胶渗透色谱和反相高效液相色谱柱分离过敏原。通过埃德曼降解、质谱、圆二色性、伴刀豆球蛋白A凝集素反应和抗辣根过氧化物酶多克隆抗体进行分子表征。使用免疫印迹和酶联免疫吸附测定、抑制实验以及组胺释放试验对来自对Fra e 1或Ole e 1具有已知反应性的过敏患者的血清样本以及针对Ole e 1的多克隆抗血清和单克隆抗体进行免疫表征。用作参考的蛋白质是在毕赤酵母中产生的rFra e 1。
纯化的nFra e 1表现为5种不同糖基化程度的变体。从使用圆二色性的光谱分析推断，nFra e 1和rFra e 1的折叠方式相同。纯化后，两种分子共享抗原性和过敏原性表位，nFra e 1的聚糖基团是潜在表位。天然Fra e 1与Ole e 1表现出强烈的交叉反应性。
天然Fra e 1是一种具有高度过敏原相关性的异质性糖基化蛋白。它与rFra e 1在结构、抗原性和过敏原性方面相似。两种蛋白质均可用于临床目的。
