Biophysical characterization of synthetic adhesins for predicting and tuning engineered living material properties

Bacterial synthetic multicellular systems are promising platforms for engineered living materials (ELMs) for medical, biosynthesis, environmental, and smart materials applications. Recent advancements in genetically encoded adhesion toolkits have enabled precise manipulation of cell-cell adhesion and the design and patterning of self-assembled multicellular materials. However, in contrast to gene regulation in synthetic biology, the characterization and control of synthetic adhesins remains limited. Here, we demonstrate the quantitative characterization of a bacterial synthetic adhesion toolbox through various biophysical methods. We determine key parameters, including number of adhesins per cell, in-membrane diffusion constant, production and decay rates, and bond-breaking force between adhesins. With these parameters, we demonstrate the bottom-up prediction and quantitative tuning of macroscopic ELM properties (tensile strength) and, furthermore, that cells inside ELMs are connected only by a small fraction of available adhesins. These results enable the rational engineering, characterization, and modeling of other synthetic and natural adhesins and multicellular consortia.