Supplementary Material for: Recombinant Fusion Proteins Assembling Der p 1 and Der p 2 Allergens from <i>Dermatophagoides pteronyssinus</i> Bussières L. Bordas-Le Floch V. Bulder I. Chabre H. Nony E. Lautrette A. Berrouet C. Nguefeu Y. Horiot S. Baron-Bodo V. 10.6084/m9.figshare.5121151.v1 https://karger.figshare.com/articles/dataset/Supplementary_Material_for_Recombinant_Fusion_Proteins_Assembling_Der_p_1_and_Der_p_2_Allergens_from_i_Dermatophagoides_pteronyssinus_i_/5121151 <i>Background:</i> Fusion proteins assembling multiple allergens can be engineered by recombinant DNA technologies in order to produce tools for diagnostic and immunotherapeutic purposes. Herein, we developed and characterized chimeras assembling Der p 1 and Der p 2 allergens as potential candidate vaccines against house dust mite allergy. <i>Methods:</i> Fusion proteins encompassing Der p 2 with either mature or proDer p 1 were expressed in <i>Escherichia coli</i> or <i>Pichia pastoris</i>. Forms with mutation in Der p 1 catalytic site were also engineered. Purified chimeras were characterized by immunoblotting, circular dichroism, disulfide bond mapping, basophil and T lymphocyte stimulation assays. <i>Results:</i> Four fusion proteins were expressed in <i>E. coli </i>as inclusion bodies, whereas only chimeras comprising proDer p 1 were obtained in yeast. All such hybrids formed polymers and aggregates, and yeast-expressed chimeras were unstable. Circular dichroism analysis performed after refolding of bacteria expressed chimeras encompassing mature Der p 1 confirmed partial folding, consistent with the occurrence of both correct and inappropriate intramolecular disulfide bonds. All fusion molecules were recognized by Der p 1- and Der p 2-specific human IgEs, monoclonal and polyclonal antibodies. Fusion proteins activate basophils from mite-allergic patients and trigger the proliferation of specific CD4+ T cells, albeit to a lower level when compared to individual allergens. <i>Conclusions:</i> Production of multiple Der p 1-Der p 2 fusion proteins exhibiting partial folding and proper antigenic properties has been achieved. Nonetheless, significant solubility and stability issues currently limit the application of such chimeras for immunotherapy or diagnostic. 2010-04-21 00:00:00 Allergy vaccine Der p 1 Der p 2 Fusion protein House dust mite Immunotherapy Recombinant allergen