Expression of the house dust mite allergen Der p 2 in the baker's yeast Saccharomyces cerevisiae

Authors

  • Hakkaart,

    1. Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and the Laboratory for Experimental and Clinical Immunology of the University of Amsterdam,
    2. Section for Molecular Biology, Department of Molecular Cell Biology, Biocentre Amsterdam, University of Amsterdam,
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  • Harmsen,

    1. Institute for Animal Science and Health, Lelystad, The Netherlands,
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  • Chua,

    1. Graduate Institute of Immunology, National Taiwan University College of Medicine, Tapei, Taiwan, Republic of China,
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  • Thomas,

    1. The Western Australian Research Institute for Child Health, Perth, Western Australia
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  • Aalberse,

    1. Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and the Laboratory for Experimental and Clinical Immunology of the University of Amsterdam,
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  • Van Ree

    1. Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and the Laboratory for Experimental and Clinical Immunology of the University of Amsterdam,
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van Ree Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and the Laboratory for Experimental and Clinical Immunology of the University of Amsterdam, Plesmanlaan 125, 1066 CX Amsterdam, The Netherlands.

Abstract

Background and Results

The major house dust mite allergen Der p 2 was expressed as a recombinant mature protein in the baker's yeast Saccharomyces cerevisiae. The yeast produces the protein fused to the invertase signal peptide, leading to the secretion of Der p 2 as a soluble protein into the culture medium. The signal peptide is hereby cleaved off, resulting in a mature allergen. In this system Der p 2 was produced in 7.6 (± 2.9) mg/L growth culture. Purification of the recombinant allergen was achieved by a single gel filtration step, resulting in a purity ≥ 95%. The yeast-derived Der p 2 was almost indistinguishable from natural Der p 2 with respect to IgE-reactivity and binding to the majority of Der p 2 specific MoAbs — as was shown in RAST analysis (n = 168) and a sandwich ELISA and RIA analysis, respectively. Recombinant and natural Der p 2 also showed similar biological activity in histamine release assays (n = 4).

Conclusion

An expression system for Der p 2 was developed that enables the production of a soluble allergen in the culture supernatant with immunological characteristics similar to the natural allergen. In addition, yeast offers the advantage of the absence of endotoxin in comparison to E. coli. This might facilitate acceptance of recombinant allergens for in vivo applications as immunotherapy or skin-prick testing.

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