High-level expression and purification of the major house dust mite allergen Der p 2 in Escherichia coli

Abstract

Der p 2, a major allergen derived from the house dust mite Dermatophagoides pteronyssinus, is one of the most clinically relevant allergens worldwide. Recombinant Der p 2 (rDer p 2) is useful in clinical diagnosis and disease-specific immunotherapy. However, previous studies showed that Der p 2 can only be expressed in Escherichia coli (E. coli) cells as inclusion bodies, thus protein refolding is required to obtain functional products. Here we report a new method to produce biologically active Der p 2 protein in E. coli. N-terminal hexahistidine- and trigger factor (TF)-tagged Der p 2 was expressed in soluble form in E. coli and purified using a combination of chromatography processes. This procedure produced milligram-level high purity Der p 2 per liter of bacterial culture. Moreover, far-UV region circular dichroism (CD) analysis and serum specific IgE reactivity test demonstrated that the secondary structure and IgE reactivity properties of rDer p 2 produced in our study were almost identical to those of natural Der p 2 (nDer p 2). In conclusion, the method developed in this work provides a useful tool for the production of immunologically active recombinant Der p 2 for clinical applications.

Keywords: Allergy; Der p 2; House dust mite; Recombinant protein.