ANALYSIS OF SEQUENCE POLYMORPHISM OF A MAJOR MITE ALLERGEN, DER-P-2

Background The major house dust mite allergen Der p 2 has been regarde d as an important allergen involved in the immunopathogenesis of aller gic asthma and eczema. Objectives To determine the degree of sequence polymorphism exists in Der p 2 at both the genomic DNA and cDNA levels . Methods Isolation of cDNA clones was performed by screening the cDNA libraries with Der p 2 cDNA probe and the genomic sequences for Der p 2 were obtained by PCR amplification from environmental dust mites wi th Der p 2-specific primers. DNA sequencing was carried out by the did eoxynucleotide chain termination method. The study of Der p 2 isoforms was performed by 2-D gel immunoblot analysis using mouse anti-rer p 2 serum. Results In this study, we have characterized the sequences of three different gene alleles coding for the major house dust mite alle rgen Der p 2 at the cDNA level. The translated polypeptides from these clones differed from each other by 3-4 amino-acid residues. These pol ymorphic residues determined were also found in Der f 2 and they were located in regions containing T-epitopes. In addition, the genomic DNA sequences of Der p 2 which were obtained by PCR amplification using t he environmental mites isolated from Taiwan and Australia have helped to confirm the authenticity of the polymorphisms detected in the cDNA clones generated from CSL cultured mites. Furthermore, 2D- immunoblot analysis indicated that there were at least 10 different isoforms (p 1 values range from greater than 7.0-5.9) of Der p 2 proteins produced by CSL cultured mites. Conclusion The results showed that there was a small but significant degree of sequence polymorphisms exists in Der p 2 gene alleles. Interestingly, the polymorphic residues were found in regions containing previously determined T-epitopes. The polymorphism data reported here will be important for the understanding of the imm une responses of mite allergens as well as for the development of the peptide-based immunotherapeutic reagents for mite allergy.