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Dual display of proteins on the yeast cell surface simplifies quantification of binding interactions and enzymatic bioconjugation reactions
Author(s) -
Lim Sungwon,
Glasgow Jeff E.,
Filsinger Interrante Maria,
Storm Erica M.,
Cochran Jennifer R.
Publication year - 2017
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.201600696
Subject(s) - yeast , bioconjugation , fusion protein , epitope , phage display , saccharomyces cerevisiae , biochemistry , protein engineering , peptide , peptide library , expression vector , biology , enzyme , green fluorescent protein , microbiology and biotechnology , chemistry , peptide sequence , recombinant dna , antibody , genetics , gene
Yeast surface display, a well‐established technology for protein analysis and engineering, involves expressing a protein of interest as a genetic fusion to either the N‐ or C‐terminus of the yeast Aga2p mating protein. Historically, yeast‐displayed protein variants are flanked by peptide epitope tags that enable flow cytometric measurement of construct expression using fluorescent primary or secondary antibodies. Here, we built upon this technology to develop a new yeast display strategy that comprises fusion of two different proteins to Aga2p, one to the N‐terminus and one to the C‐terminus. This approach allows an antibody fragment, ligand, or receptor to be directly coupled to expression of a fluorescent protein readout, eliminating the need for antibody‐staining of epitope tags to quantify yeast protein expression levels. We show that this system simplifies quantification of protein‐protein binding interactions measured on the yeast cell surface. Moreover, we show that this system facilitates co‐expression of a bioconjugation enzyme and its corresponding peptide substrate on the same Aga2p construct, enabling enzyme expression and catalytic activity to be measured on the surface of yeast.