Biophysical Characterization of Synthetic Adhesins for Developing Tunable Engineered Living Materials With Predictive Properties

Abstract

The emerging discipline of engineered living materials (ELMs) highlights many outstanding properties over traditional materials, and synthetic biology approaches are considered a key up-and-coming approach for ELM. While synthetic adhesins have been developed in other systems, recent advancements in a genetically encoded adhesion toolkit for E. coli have enabled precise manipulation of cell-cell adhesion and the design of self-assembled multicellular patterns and materials. However, while synthetic gene regulation in synthetic biology is well described, the characterization of synthetic adhesins remains limited, hindering their functionality and the programming of ELMs. We demonstrate on example of one particular adhesion toolbox how it can be characterized in general and we report quantitative measures of key biophysical parameters, including the total number of adhesion proteins per cell, lateral membrane diffusion, production and degradation rates, and binding force. For example, the viscoelasticity of an ELM depends on whether cells may slide past each other and rearrange their relative position and orientation without the need to break adhesion bonds, critically depends on the number of adhesins per cell, their stability (longevity), their pairwise binding strength, and their ability to move within the cellular membrane. Based on these measured parameters, we then demonstrate how to predict bottom-up tunable macroscopic ELM properties with a focus on material tensile strength as a suitable example that is relevant, for example, for bioprinting. Furthermore, we show how small molecular changes affect the viscosity of the material and we aim to provide a systematic quantification of these effects for the future development of machine-learning algorithms with strong predictive capabilities for rational material engineering.