Abstract:
Recent identified Der p 23 has been recognized as an important house dust mite (HDM) allergen which displays high IgE reactivity comparable to the major HDM allergens Der p 1 and Der p 2. Due to limited amounts of the natural allergen, the in-depth characterization of Der p 23 requires the production of a recombinant form. Present study aims were to produce a recombinant form of Der p 23 using P. pastoris expression system subsequently characterize its physicochemical properties and to analyze for the first time the IgE binding frequency of Der p 23 in a cohort of Thai HDM allergic patients as well as to evaluate the ability to activate innate immunity. The expression of secreted mature rDer p 23 reaches maximum at 48 hrs under 2% methanol induction and resulted in N-terminal truncation with longer induction period. rDer p 23 is characterize as a mannosylated protein (possibly modified at amino acid T30-T32) with two intra-disulfide bonds and adopt mainly unfolded structure. Polyclonal antibodies to rDer p 23 can detect the natural allergen in aqueous fecal pellets extracts suggesting that both forms of Der p 23 share common B-cell epitopes. rDer p 23 as well as the natural corresponding allergen were unable to interact in-vitro with chitin matrices. More than fifty percent of Thai HDM allergic patients (n=222) developed Der p 23-specific IgE which was comparable with the IgE binding frequency of rDer p 2 (67%). Meanwhile, the allergenicity of rDer p 23 was highlighted through degranulation of RBL cells expressing human FcεRI. Nevertheless, human airway epithelial cells were able to produce IL-8 in response to rDer p 23 (5-20µg/ml) through NF-κB and MAPK (MEK, JNK, and p38)-dependent activation signaling pathways. Our findings highlighted important levels of Der p 23 sensitizations in Thailand. The protease sensitivity of Der p 23 as well as the very limited release of this allergen from extracted mite fecal pellets clearly suggested that rDer p 23 may be a valuable material to replace standardized HDM extracts for the diagnosis of Der p 23 sensitivities.
