Arruda L K, Vailes L D, Hayden M L, Benjamin D C, Chapman M D
Department of Internal Medicine, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
J Biol Chem. 1995 Dec 29;270(52):31196-201. doi: 10.1074/jbc.270.52.31196.
An allergen cloned from a Blattella germanica (German cockroach) cDNA library, encoded a 182-amino acid protein of 20,904 Da. This protein, designated B. germanica allergen 4 (Bla g 4), was expressed as a glutathione S-transferase fusion protein in Escherichia coli and purified by affinity chromatography and high-performance liquid chromatography. The prevalence of serum IgE antibody to recombinant Bla g 4 in 73 cockroach allergic patients with asthma ranged from 40% (antigen binding radioimmunoassay) to 60% (plaque immunoassay). Cockroach allergic patients gave positive intradermal skin tests to recombinant Bla g 4 at concentrations of 10(-3)-10(-5) micrograms/ml, whereas non-allergic controls, or cockroach allergic patients with no detectable serum IgE antibody to Bla g 4, gave negative skin tests to 1 microgram/ml. Polymerase chain reaction and Southern analysis identified a 523-base pair DNA encoding Bla g 4 in both B. germanica and Periplaneta americana (American cockroach). However, Northern analysis showed that mRNA encoding Bla g 4 was transcribed in B. germanica but not in P. americana, suggesting that allergen expression was species specific. Sequence similarity searches showed that Bla g 4 was a ligand binding protein or calycin and unexpectedly revealed that this family contained several important allergens: beta-lactoglobulin, from cow milk, and rat and mouse urinary proteins. Although the overall sequence homology between these proteins was low (approximately 20%), macromolecular modeling techniques were used to generate two models of the tertiary structure of Bla g 4, based on comparisons with the x-ray crystal coordinates of bilin binding protein and rodent urinary proteins. The results show that members of the calycin protein family can cause IgE antibody responses by inhalation or ingestion and are associated with asthma and food hypersensitivity.
从德国小蠊（德国蟑螂）cDNA文库中克隆出的一种变应原，编码一种分子量为20,904 Da的182个氨基酸的蛋白质。这种蛋白质被命名为德国小蠊变应原4（Bla g 4），它在大肠杆菌中作为谷胱甘肽S-转移酶融合蛋白表达，并通过亲和层析和高效液相色谱法进行纯化。在73名患有哮喘的蟑螂过敏患者中，针对重组Bla g 4的血清IgE抗体阳性率在40%（抗原结合放射免疫测定法）至60%（噬斑免疫测定法）之间。蟑螂过敏患者对浓度为10(-3)-10(-5)微克/毫升的重组Bla g 4进行皮内皮肤试验呈阳性，而无过敏反应的对照者，或对Bla g 4无血清IgE抗体检测到的蟑螂过敏患者，对1微克/毫升的试验呈阴性。聚合酶链反应和Southern分析在德国小蠊和美洲大蠊（美国蟑螂）中均鉴定出一个编码Bla g 4的523个碱基对的DNA。然而，Northern分析表明，编码Bla g 4的mRNA在德国小蠊中转录，而在美洲大蠊中未转录，这表明变应原表达具有物种特异性。序列相似性搜索显示，Bla g 4是一种配体结合蛋白或亲环素，并且意外地发现该家族包含几种重要的变应原：来自牛奶的β-乳球蛋白以及大鼠和小鼠的尿液蛋白。尽管这些蛋白质之间的总体序列同源性较低（约20%），但基于与胆汁素结合蛋白和啮齿动物尿液蛋白的X射线晶体坐标进行比较，利用大分子建模技术生成了Bla g 4三级结构的两种模型。结果表明，亲环素蛋白家族成员可通过吸入或摄入引发IgE抗体反应，并与哮喘和食物超敏反应相关。
