Batanero E, Gonzalez De La Peña M A, Villalba M, Monsalve R I, Martin-Esteban M, Rodríguez R
Departamento de Bioquímica y Biología Molecular, Facultad de Química, Universidad Complutense, Madrid, Spain.
Clin Exp Allergy. 1996 Dec;26(12):1401-10.
An olive allergen-like protein has been detected in privet pollen. This protein could be involved in the allergenic cross-reactivity described for privet and olive tree pollen extracts.
Isolation and characterization of natural Lig v 1. Cloning and expression of its cDNA in order to assess its structural similarity with the olive allergen.
Current chromatographic methods were used to isolate the privet counterpart of Ole e 1. A pool of sera from subjects allergic to olive tree pollen was used to immunodetect the protein in the elution profiles. Ole e 1-specific polyclonal antibody and allergic sera were used in immunoblotting assays of the isolated protein. Polymerase chain reaction amplification of the first strand cDNA synthesized from the privet pollen total RNA was carried out to prepare a full-length fragment encoding Lig v 1. After nucleotide sequencing, expression of one clone was performed in Escherichia coli, under the form of a fusion protein with glutathione S-transferase. The IgE binding capability of the recombinant protein was also analysed.
The major allergen from privet pollen. Lig v 1, was purified to homogeneity by two gel filtration chromatographies and one reverse-phase high-performance liquid chromatography. Its amino acid composition and N-terminal amino acid sequence were determined. Two different clones encoding Lig v 1 were sequenced. Strong sequence similarity between Lig v 1 and Ole e 1 was observed, the identity being 85 and 96%. One of the sequenced clones was expressed and the recombinant product exhibited IgG and IgE binding activities against both anti-Ole e 1 polyclonal antibodies and olive-allergic sera.
Privet pollen contains a protein structurally and immunologically related to the major allergen of olive pollen. The similarity exhibited by these proteins could explain the cross-reactivity observed between the two pollen extracts. Since these allergens are highly polymorphic, the expression of an immunologically active recombinant Lig v 1 will permit the preparation of well defined molecules for both research and clinical purposes.
在女贞花粉中检测到一种类似橄榄过敏原的蛋白质。该蛋白质可能参与了女贞和橄榄树花粉提取物之间描述的过敏交叉反应。
天然Lig v 1的分离与鉴定。克隆其cDNA并进行表达，以评估其与橄榄过敏原的结构相似性。
采用现行色谱方法分离女贞中与Ole e 1相对应的蛋白。使用对橄榄树花粉过敏的受试者的血清池对洗脱图谱中的蛋白质进行免疫检测。将Ole e 1特异性多克隆抗体和过敏血清用于分离蛋白的免疫印迹分析。从女贞花粉总RNA合成的第一链cDNA进行聚合酶链反应扩增，以制备编码Lig v 1的全长片段。核苷酸测序后，在大肠杆菌中以与谷胱甘肽S-转移酶融合蛋白的形式对一个克隆进行表达。还分析了重组蛋白的IgE结合能力。
通过两次凝胶过滤色谱和一次反相高效液相色谱将女贞花粉中的主要过敏原Lig v 1纯化至同质。确定了其氨基酸组成和N端氨基酸序列。对编码Lig v 1的两个不同克隆进行了测序。观察到Lig v 1与Ole e 1之间有很强的序列相似性，同一性分别为85%和96%。对其中一个测序克隆进行了表达，重组产物对两种抗Ole e 1多克隆抗体和橄榄过敏血清均表现出IgG和IgE结合活性。
女贞花粉含有一种在结构和免疫上与橄榄花粉主要过敏原相关的蛋白质。这些蛋白质表现出的相似性可以解释两种花粉提取物之间观察到的交叉反应。由于这些过敏原具有高度多态性，具有免疫活性的重组Lig v 1的表达将允许制备用于研究和临床目的的明确分子。
