biostudies-literatureLei MNational Institutes of HealthNIGMS NIH HHS399-408https://www.ebi.ac.uk/biostudies/studies/S-EPMC10072783biostudies-literatureUnknown120(2)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.Biotechnology and bioengineeringFlow cytometric evaluation of yeast-bacterial cell-cell interactions.PMC10072783R35 GM133471Lei MNair NUTrivedi VDLee KVan Deventer JAfalseFlow cytometric evaluation of yeast-bacterial cell-cell interactions.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.2023-01-01T00:00:00Z2023 Feb2025-07-12T03:04:25.38Z2025-04-06T13:24:00.114ZS-EPMC100727833625911010.1002/bit.28253