Cloning and Characterisation of Two IgE-Binding Proteins, Homologous to Tropomyosin and α-Tubulin, from the Mite <i>Lepidoglyphus destructor</i>

Abstract

BACKGROUND: The dust mite Lepidoglyphus destructor is a major source of mite allergy in European rural environments, but it also causes allergy in urban populations around the world. We have previously cloned, sequenced and expressed several allergens from L. destructor (Lep d 2, Lep d 5, Lep d 7 and Lep d 13). The aim of this study was to identify and clone additional allergens from L. destructor, and to evaluate their IgE-binding reactivities. METHODS: PCR and screening with sera from L. destructor-sensitised individuals were used to isolate new clones from a phage display L. destructor cDNA library. The complete coding sequences of the clones were determined and expressed as His(6)-tagged recombinant proteins in Escherichia coli. The recombinant proteins were analysed by SDS-PAGE, immunoblotting and mass spectrometry. RESULTS: Two new clones, showing homology to tropomyosin and alpha-tubulin in several species, were isolated from the phage display L. destructor cDNA library. Due to its homology to group 10 dust mite allergens, the tropomyosin clone was named Lep d 10. The IgE-binding frequencies of the recombinant Lep d 10 and alpha-tubulin were 13% (18/136) and 12% (11/95), respectively, among subjects with IgE reactivity to mites and/or crustaceans. CONCLUSIONS: Two new allergens from L. destructor have been identified and can now be added to the repertoire of recombinant L. destructor allergens. In addition, both these allergens belong to highly conserved protein families and may be important for evaluation of allergenic cross-reactivity.