Anais da Academia Brasileira de Ciências
Print version ISSN 0001-3765
YU-BAO, Cui et al. Cloning, expression, and analysis of the group 2 allergen from Dermatophagoides farinae from China. An. Acad. Bras. Ciênc. [online]. 2010, vol.82, n.4, pp. 941-951. ISSN 0001-3765. http://dx.doi.org/10.1590/S0001-37652010000400017.
To obtain the recombinant group 2 allergen product of Dermatophagoides farinae (Der f 2), the Der f 2 gene was synthesized by RT-PCR. The full-length cDNA comprised 441 nucleotides and was 99.3% identical to the reference sequence (GenBank AB195580). The cDNA was bound to vector pET28a to construct plasmid pET28a(+)-Der f 2, which was transformed into E. coli BL21 and induced by IPTG. SDS-PAGE showed a specific band of about 14kDa in the hole cell lysate. s estiated by chroatography, about 3.86 g of the recobinant product as obtained, which conjugated with serum IgE from asthmatic children. The protein had a signal peptide of 17 amino acids. Its secondary structure comprised an alpha helix (19.86%), an extended strand (30.82%), and a random coil (49.32%). The subcellular localization of this allergen was predicted to be at mitochondria. Furthermore, its function was shown to be associated with an MD-2-related lipid-recognition (ML) domain. The results of this study provide a solid foundation for large-scale production of the allergen for clinical diagnosis and treatent of allergic disorders.
Keywords : Dermatophagoides farinae; house-dust-mite allergy; mite allergens; Der f 2; allergen engineering; immunotherapy; bioinformatics.