Design of temperature-responsive biofilms for the precise and reversible control of engineered living biomaterials

• Applicant: Dr. Jan Mathony
• Subject Area: Biomaterials ; Cell Biology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 520612620

Biomaterials that combine certain mechanical properties with the versatility of living systems, are becoming increasingly important in response to the ecological and technological challenges of our time. A promising starting point for such "engineered living materials" (ELMs) are bacterial biofilms, such as the "curli" system of Escherichia coli. Such biofilms can be sustainably produced and processed into macroscopic materials such as hydrogels or bioplastic. Importantly, curli materials exhibit high mechanical stability and resistance to harsh chemical conditions. Curli themselves consist of fibrils that are formed by polymerization of the protein CsgA secreted by E. coli and are then anchored to the cell surface by its binding partner CsgB. Although the formation of biofilms is in general genetically inducible, the mechanical properties of existing curli biofilms can hardly be changed after their formation due to the high stability of the fibrils. Thus, the mechanical adaptation of the biofilm to external stimuli has thus far not been possible. This project aims to develop molecular tools for reversible control of the mechanical stability of curli-based ELMs. Using modern protein engineering methods, I will generate artificial, temperature-sensitive CsgA variants. I will then use these to control the biophysical properties of curli fibrils in response to temperature changes. In the first part of the project, I will generate fusion proteins consisting of CsgA and various receptor domains, which homo-dimerize depending on the temperature and can thus form stabilizing, reversible cross-connections between individual fibrils. This makes it possible to specifically control the rheological properties of the resulting ELMs. In a second step, I will finally produce synthetic CsgA variants that only oligomerize at low temperatures (<25°C) by combining CsgA point mutation libraries with domain insertion methods. I will use these temperature-sensitive, reversible and adaptive curli biofilms in subsequent projects together with collaboration partners for 3D bioprinting of living structures and for the programmed release of probiotic bacteria. By applying protein engineering to curli biofilms, this research project will unlock a new generation of biomaterials.

• DFG Programme: Research Grants