Recombinant Fusion Proteins Assembling Der p 1 and Der p 2 Allergens from Dermatophagoides pteronyssinus

被引：16

作者：

Bussieres, Laetitia ^{[1
]}

Bordas-Le Floch, Veronique ^{[1
]}

Bulder, Ingrid ^{[1
]}

Chabre, Henri ^{[1
]}

Nony, Emmanuel ^{[1
]}

Lautrette, Aurelie ^{[1
]}

Berrouet, Christelle ^{[1
]}

Nguefeu, Yvette ^{[1
]}

Horiot, Stephane ^{[1
]}

Baron-Bodo, Veronique ^{[1
]}

Van Overtvelt, Laurence ^{[1
]}

De Conti, Anne Marie ^{[1
]}

Schlegel, Anne ^{[2
]}

Maguet, Nicolas ^{[2
]}

Mouz, Nicolas ^{[2
]}

Lemoine, Pierrick ^{[1
]}

Batard, Thierry ^{[1
]}

Moingeon, Philippe ^{[1
]}

机构：

[1] Stallergenes, Res & Dev, Antony, France

[2] PX Therapeut, Grenoble, France

来源：

INTERNATIONAL ARCHIVES OF ALLERGY AND IMMUNOLOGY | 2010年 / 153卷 / 02期

关键词：

Allergy vaccine; Der p 1; Der p 2; Fusion protein; House dust mite; Immunotherapy; Recombinant allergen; HOUSE-DUST MITE; HIGH-LEVEL EXPRESSION; MAJOR ALLERGEN; CRYSTAL-STRUCTURE; PICHIA-PASTORIS; IGE; IMMUNOTHERAPY; BINDING; FARINAE; POLLEN;

D O I：

10.1159/000312631

中图分类号：

R392 [医学免疫学];

学科分类号：

100102 ;

摘要：

Background: 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. Methods: Fusion proteins encompassing Der p 2 with either mature or proDer p 1 were expressed in Escherichia coli or Pichia pastoris. 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. Results: Four fusion proteins were expressed in E. coli 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. Conclusions: 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. Copyright (C) 2010 S. Karger AG, Basel

引用

页码：141 / 151

页数：11

共 35 条

[1] Engineering of major house dust mite allergens Der p 1 and Der p 2 for allergen-specific immunotherapy [J].