The production of allergens from cDNA clones will provide a clonally pure source of material for experimental and perhaps clinical studies. Attempts to produce the major mite allergen, Der p I, in a highly antigenic form in bacteria have, to date, had limited success. In this study, a high level of production of Der p I from a Cup1 gene cassette from pYELC5-13T in Saccharomyces cerevisiae is described. Although the protein was insoluble, it could be readily solubilized in a urea solution and remained in solution when it was returned to more physiologic buffers. An amount equivalent to about 1 mg/L of yeast culture could then be isolated by affinity chromatography with an immobilized monoclonal antibody. This product reacted strongly with IgE in 9/11 sera from mite-allergic patients compared to the 50% reactivity achieved for Der p I previously produced as a fusion by bacteria. Similarly, the intensity of binding and ability to absorb out Der p I specificities were much greater for the yeast, pYELC5-13T, product. Studies with monoclonal antibodies also demonstrated the yeast, Der p I, had a high degree of antigenicity, although clear differences with the native allergen were demonstrated. The high frequency of reactivity with IgE of the pYELC5-13T formally demonstrates that a single gene product of Der p I is a major allergen and demonstrates that even for Der p I, which is synthesized from a proenzyme, considerable antigenicity can be obtained by expressing the mature protein.

• PMID: 1730842
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• PubMed
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Reference Type

Journal Article | Research Support, Non-U.S. Gov't

Authors

Chua KY, Kehal PK, Thomas WR, Vaughan PR, Macreadie IG

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