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bioRxiv · 10.1101/2023.06.30.547162

Non-covalent Linking Antibody to Yeast Using the Cell Surface Display of Staphylococcal Protein A

Shi, Y.-F.
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Abstract

Yeast surface display represents a commonly used platform suitable for the generation and screening of antibodies as well as the selection of high-producer clones. The methods of yeast display rely on the genetic fusion of recombinant antibodies to an abundant cell wall protein of yeast. Here, the study of proof of concept showed that the conventional strategy of expression of fusion antibodies was replaced by non-covalent binding to antibodies for yeast surface display. The use of cell surface display of an epitope tag will endow the cells with new arms for immobilizing, absorbing or targeting the proteins. Staphylococcal protein A, which is characterized by its ability to bind selectively to the Fc region of IgG, was examined to be expressed on the surface of Saccharomyces cerevisiae using a secretion signal of Rhizopus oryzae glucoamylase and C-terminal half of -agglutinin including glycosylphosphatidylinositol (GPI) anchor attachment signal under the control of the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) promoter. On the other hand, an Fc-fused enzyme was created to construct a molecular fusion of Rhizopus oryzae Lipase with the spacer and the Fc region of IgG heavy chain. The secretion of fusion protein was carried out using pre--factor leader region as secretion signal under the control of the 5-upstream region of the Candida tropicalis isocitrate lyase gene (UPR-ICL) in S. cerevisiae. The Fc-fused lipase was captured by Staphylococcal protein A as an adaptor protein displayed on the surface of yeast cells. The method of this switchable yeast display takes advantage of the "secretion-and-capture" strategy and can be applied to improve the efficiency of yeast display of full-length IgG. graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/547162v1_ufig1.gif" ALT="Figure 1"> View larger version (88K): org.highwire.dtl.DTLVardef@1352ccforg.highwire.dtl.DTLVardef@10d5f0org.highwire.dtl.DTLVardef@14d920forg.highwire.dtl.DTLVardef@1b03e6f_HPS_FORMAT_FIGEXP M_FIG Staphylococcal protein A, which has the ability to binding to the Fc region of IgG and leaving the antigen combing site free, has been assembled on the surface of yeast cells to target antibodies or enzymes with Fc fusion. C_FIG

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Shi, Y.-F.. 2023-06-30. Non-covalent Linking Antibody to Yeast Using the Cell Surface Display of Staphylococcal Protein A. https://doi.org/10.1101/2023.06.30.547162

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