Cloning, expression, and analysis of the group 2 allergen from Dermatophagoides farinae from China

• Autor(a) principal: Yu-bao,Cui
• Data de Publicação: 2010
• Outros Autores: Zhou,Ying, Weihong,Shi, Guifang,Ma, Yang,Li, Yungang,Wang
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Anais da Academia Brasileira de Ciências (Online)
• Texto Completo: http://old.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652010000400017
• topic: Dermatophagoides farinae; house-dust-mite allergy; mite allergens; Der f 2; allergen engineering; immunotherapy; bioinformatics

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.