Gene synthesizing, expression and immunogenicity characterization of recombinant translation elongation factor 2 from Dermatophagoides farinae

Abstract

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, 785-790,811-817and827-836)and14peptidesasTcellepitopes (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.