Flow cytometric evaluation of yeast–bacterial cell–cell interactions

Abstract

Synthetic cell-cell interaction systems can be useful for understanding multicellular communities or for screening binding molecules. We adapt a previously characterized set of synthetic cognate nanobody-antigen pairs to a yeast-bacteria coincubation format and use flow cytometry to evaluate cell-cell interactions mediated by binding between surface-displayed molecules. We further use fluorescence-activated cell sorting to enrich a specific yeast-displayed nanobody within a mixed yeast-display population. Finally, we demonstrate that this system supports the characterization of a therapeutically relevant nanobody-antigen interaction: a previously discovered nanobody that binds to the intimin protein expressed on the surface of enterohemorrhagic Escherichia coli. Overall, our findings indicate that the yeast-bacteria format supports efficient evaluation of ligand-target interactions. With further development, this format may facilitate systematic characterization and high-throughput discovery of bacterial surface-binding molecules.

Small Antibodies with Big Applications: Nanobody-Based Cancer Diagnostics and Therapeutics

“…As a eukaryotic expression system, it can display larger and more complex proteins with posttranslational modifications. Furthermore, yeast is large enough to facilitate the screening of the desired nanobodies using advanced techniques, such as flow cytometry or magnetic bead sorting methods [21,22]. For example, Chen et al developed a SpyTag/SpyCatcher cell-surface protein-binding system that differs from traditional yeast display techniques.…”