State Key Laboratory of Respiratory Disease for Allergy at Shenzhen University, Shenzhen Key Laboratory of Allergy and Immunology, Shenzhen University School of Medicine, Shenzhen, Guangdong 518060, P.R. China.
Department of Allergy, Affiliated Luohu Hospital of Shenzhen University, Shenzhen Luohu Hospital Group, Shenzhen, Guangdong 518000, P.R. China.
Mol Med Rep. 2019 Dec;20(6):5324-5334. doi: 10.3892/mmr.2019.10786. Epub 2019 Oct 31.
House dust mite (HDM) hypersensitivity increasingly affects millions of individuals worldwide. Although numerous major allergens produced by HDM species have been characterized, some of the less potent allergens remain to be studied. The present study aimed to obtain the recombinant allergen of Translation Elongation Factor 2 (TEF 2) from the HDM Dermatophagoides farinae by synthesizing, and then expressing the recombinant TEF 2 to identify its immunogenicity. In the present study, the D. farinae TEF 2 (Der f TEF 2) was synthesized, expressed and purified. The molecular characteristics of Der f TEF 2 were analyzed using bioinformatics approaches. The recombinant protein was purified via affinity chromatography, and the allergenicity was assessed using immunoblotting, ELISAs and skin prick tests. The gene for TEF 2 consists of 2,535 bp and encodes an 844‑amino acid protein. A positive response to recombinant Der f TEF 2 was detected in 16.2% of 37 patients with HDM allergies using skin prick tests. In addition, the immunoblotting indicated that the protein showed a high ability to bind serum IgE from patients allergic to HDMs, and that the recombinant TEF 2 was highly immunogenic. Bioinformatics analysis predicted 17 peptides as B cell epitopes (amino acids 29‑35, 55‑64, 92‑99, 173‑200, 259‑272, 311‑318, 360‑365, 388‑395, 422‑428, 496‑502, 512‑518, 567‑572, 580‑586, 602‑617, 785‑790, 811‑817 and 827‑836) and 14 peptides as T cell epitopes (amino acids 1‑15, 65‑79, 120‑134, 144‑159, 236‑250, 275‑289, 404‑418, 426‑440, 463‑477, 510‑524, 644‑658, 684‑698, 716‑730 and 816‑830). The software DNAStar predicted the secondary structure of TEF 2, and showed that 27 α‑helices and five β‑sheets were found in the protein. In conclusion, the present study cloned and expressed the Der f TEF 2 gene, and the recombinant protein exhibited immunogenicity, providing a theoretical bases, and references, for the diagnosis and treatment of allergic disease.
屋尘螨（HDM）过敏症日益影响着全球数以百万计的人群。尽管已经对 HDM 物种产生的许多主要过敏原进行了描述，但一些较弱的过敏原仍有待研究。本研究旨在通过合成和表达重组翻译延伸因子 2（TEF 2），获得屋尘螨的重组过敏原，以鉴定其免疫原性。
在本研究中，合成、表达和纯化了屋尘螨的 TEF 2（Der f TEF 2）。使用生物信息学方法分析 Der f TEF 2 的分子特征。通过亲和层析纯化重组蛋白，并通过免疫印迹、ELISA 和皮肤点刺试验评估其变应原性。TEF 2 基因由 2535 bp 组成，编码 844 个氨基酸的蛋白质。皮肤点刺试验显示，37 例 HDM 过敏患者中有 16.2%对重组 Der f TEF 2 呈阳性反应。此外，免疫印迹表明，该蛋白具有与对 HDM 过敏的患者血清 IgE 结合的高能力，并且该重组 TEF 2 具有高度的免疫原性。生物信息学分析预测了 17 个肽段作为 B 细胞表位（氨基酸 29-35、55-64、92-99、173-200、259-272、311-318、360-365、388-395、422-428、496-502、512-518、567-572、580-586、602-617、785-790、811-817 和 827-836）和 14 个肽段作为 T 细胞表位（氨基酸 1-15、65-79、120-134、144-159、236-250、275-289、404-418、426-440、463-477、510-524、644-658、684-698、716-730 和 816-830）。软件 DNAStar 预测了 TEF 2 的二级结构，显示该蛋白中存在 27 个α-螺旋和 5 个β-折叠。
综上所述，本研究克隆并表达了 Der f TEF 2 基因，该重组蛋白具有免疫原性，为过敏性疾病的诊断和治疗提供了理论基础和参考。
