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Title: Cell surface display of highly pathogenic avian influenza hemagglutinin on the surface of Pichia pastoris cells using alpha-agglutinin for production of oral vaccines

Author
item Wasilenko, Jamie
item Sarmento, Luciana
item Spatz, Stephen
item Pantin Jackwood, Mary

Submitted to: Biotechnology Progress
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/8/2009
Publication Date: 3/15/2010
Citation: Wasilenko, J.L., Sarmento, L., Spatz, S.J., Pantin Jackwood, M.J. 2010. Cell surface display of highly pathogenic avian influenza hemagglutinin on the surface of Pichia pastoris cells using alpha-agglutinin for production of oral vaccines. Biotechnology Progress. 26(2):542-547.

Interpretive Summary: Highly pathogenic avian influenza virus infections result in devastating poultry loses around the world. Traditional vaccination methods for HPAI viruses require costly and time-consuming injection of individual birds, often multiple times, in order to provide adequate protection. Furthermore methods to develop and manufacture these vaccines are time consuming and somewhat limited in the ability to generate vaccines quickly. The use of yeast as an expression system for influenza proteins can remedy these problems. Yeast glycosylate proteins more similarly to that of mammalian proteins, production of proteins in yeast is fast, inexpensive, and yeast have the added benefit of providing high quality nutrition when given in the feed. In this study, the hemagglutinin protein from a highly pathogenic avian influenza virus, subtype H5N1, was expressed on the surface of the yeast strain Pichia pastoris. Surface expression of the hemagglutinin protein was demonstrated by immunofluorescence microscopy. Functionally the expressed hemagglutinin protein retained agglutinating activity. This study represents the first step in the generation of a yeast-based vaccine for protection against highly pathogenic strains of avian influenza.

Technical Abstract: Yeast are an ideal organism to express viral antigens because yeast glycosylate proteins are more similar to mammals than bacteria, and expression of proteins in yeast is relatively fast and inexpensive. In addition to the convenience of production, for purposes of vaccination, yeast have been shown to have natural adjuvant activity making the expressed proteins more immunogenic when administered along with yeast cell wall components. Development of genetic systems to display foreign proteins on the surface of yeast via fusion to glycosylphosphatidylinositol-anchored (GPI) proteins has further simplified the production of recombinant proteins by not requiring harsh treatments for cellular lysis or protein purification. The hemagglutinin protein from a highly pathogenic avian influenza (HPAI) strain [A/Egret/HK/757.2/02], subtype H5N1, was expressed on the surface of the yeast strain Pichia pastoris, as an anchored C-terminal fusion with the Saccharomyces cerevisiae GPI-anchored cell wall protein, alpha-agglutinin. Surface expression of the hemagglutinin fusion protein was demonstrated by immunofluorescence microscopy. Functionally the fusion protein retained hemagglutinating activity. This study represents the first step in the generation of a yeast-based vaccine for protection against highly pathogenic strains of avian influenza.