Mimotopes identify conformational B-cell epitopes on the two major house dust mite allergens Der p 1 and Der p 2 (1)
House dust mites have been shown to be common indoor allergens elucidating type I hypersensitivity reactions such as asthma, perennial rhinitis and atopic dermatitis. The most important allergens are Der p 1 and Der p 2 from the species D. pteronyssinus, which are present in high concentrations in household dust and are predominantly recognized by IgE of mite allergic patients’ sera.
120 random phage clones were screened by a colony screening ELISA with each panning antibody. On this ELISA, nine phage clones showed a significantly higher bind- ing signal to the specific antibody in comparison to background reactivity with wild-type phage.
The deduced amino acid sequences of the selected mimotopes are listed. Clone 5 matched at the amino acids 11–13 and 178–183. The other part of the epitope was completed by clones 39 and 54. These two clones comprise mostly the same amino acids. They also match on an area close to the other two mimotopes, clone 22 and clone 47. All selected peptide mimotopes localized overlapping areas and thus together determined a single discontinuous structural epitope on natural Der p 2.
The linear sequence of Der p 1 and Der p 2 identified sequence stretches within the conserved regions of the allergens. Locating these matched amino acids from Der p 1 and respectively from Der p 2 on the conformational structure of other allergens within the groups, highly similar epitope patches were determined in both allergen groups.
The equal phage coating was confirmed with peroxidase-labeled anti-phage antibody. Clone 5 identified from Der p 1-biopanning showed a higher IgG and IgE recognition than the other clones. With respect to specific IgE binding clone 47 had the lowest capacity among all tested clones. Clone 25 had the highest reactivity. Via specific IgE antibodies clone 27 was recognized best.
In the structure of Der p 1, three disulfide bridges form covalent bonds between residues C4 and C117, C31 and C71, and C65 and C103. Interestingly, Der p 1 forms dimers non-covalently attached to each other. Greene et al. used linear recombinant peptides testing for IgE binding activity on immunoblot, where they found two non-overlapping peptides (53–99, 101–154) having significant binding activity.
