Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae.
BACKGROUND
Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model.


RESULTS
The proteins were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS) after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained their enzymatic activities. Phospholipase A1 severely inhibited the growth of the yeast cells. Antigen 5 - expressing yeast cells bound IgE antibodies from wasp venom allergic patient sera but not from control sera as demonstrated by FACS. Moreover, antigen 5 - expressing yeast cells were capable of mediating allergen-specific histamine release from human basophils.


CONCLUSIONS
All the three major wasp venom allergens were expressed on the yeast surface. A high-level expression, which was observed only for antigen 5, was needed for detection of IgE binding by FACS and for induction of histamine release. The non-modified S. cerevisiae cells did not cause any unspecific reaction in FACS or histamine release assay despite the expression of high-mannose oligosaccharides.In perspective the yeast surface display may be used for allergen discovery from cDNA libraries and possibly for sublingual immunotherapy as the cells can serve as good adjuvant and can be produced in large amounts at a low price.
Borodina
Irina
I
Dermato-Allergological Dept. K, CUH-Gentofte, Rigshospitalet Dept 7551, Blegdamsvej 9, 2100 København Ø, Denmark.
Jensen
Bettina M
BM
Søndergaard
Ib
I
Poulsen
Lars K
LK
D000485:Allergens / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D001491:Basophils / Q000276:immunology / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D005434:Flow Cytometry; D006632:Histamine / Q000378:metabolism; D006801:Humans; D006821:Hyaluronoglucosaminidase / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D007073:Immunoglobulin E / Q000276:immunology; D054466:Phospholipases A1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D014862:Wasp Venoms / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
C543385:AGA2 protein, S cerevisiae; D000485:Allergens; D015815:Cell Adhesion Molecules; D029701:Saccharomyces cerevisiae Proteins; D014862:Wasp Venoms; C040583:white-faced hornet venom antigen 5; D007073:Immunoglobulin E; D006632:Histamine; D054466:Phospholipases A1; D006821:Hyaluronoglucosaminidase