Correlating single-molecule rupture mechanics with cell population adhesion by yeast display

Abstract

Here, we present a method based on yeast surface display that allows for direct comparison between population-level cell adhesion strength and single-molecule receptor-ligand rupture mechanics. We developed a high-throughput yeast adhesion assay in which yeasts displaying monomeric streptavidin (mSA) or enhanced mutant mSA were adhered to a biotinylated coverglass submerged in fluid. After exposure to shear stress (20–1000 dyn/cm2) by rapid spinning of the coverglass, cells were imaged to quantify the midpoint detachment shear stress for the cell population. We then performed atomic force microscope single-molecule force spectroscopy (SMFS) on purified mSA variants and identified correlations between single-molecule rupture force distributions and cell population adhesion strength. Several features of yeast display were important for successful correlations of adhesion strength to be drawn, including covalent attachment of the receptor to the cell wall, a precisely defined molecular pulling geometry, repression of nonspecific adhesion, and control for multivalency. With these factors properly taken into account, we show that spinning disk cell adhesion assays can be correlated with SMFS and are capable of screening the mechanical strength of receptor-ligand complexes. These workflow enhancements will accelerate research on mechanostable receptor-ligand complexes and receptor-mediated cell adhesion.