Developing 3D-Hybrid Living Materials for Mixotrophic Biofilm Catalysis

• Applicants: Professorin Dr. Katja Bühler; Dr.-Ing. Rohan Karande
• Subject Area: Biological Process Engineering
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 559382400

The 3D-BioCat project is pioneering a breakthrough platform for cultivating light-powered mixotrophic biofilms to unlock sustainable production pathways for polymer precursors. Central to the project is the development of modular Hybrid Living Materials (HLMs) based reactors designed to support chemo-heterotrophic, photoautotrophic, and mixed trophic biofilms. The key hypothesis guiding the research is that photo-biofilm catalysts can be operated within 3D structures by emulating natural systems, with their productivities and scalability dependent on the reactor's design and operational parameters. To explore this hypothesis, 3D-BioCat investigates two nature-inspired HLM reactor designs: a spiral configuration leading to vertically cylindrical reactors and a semi-ellipsoid design. The project aims to evaluate both formats, focusing on two main performance indicators: catalytic efficiency in producing epsilon-caprolactone and potential scalability for industrial use. Additionally, the project will assess and enhance the long-term catalytic efficiency of recombinant biofilms in sustainable epsilon-caprolactone production. Furthermore, an innovative scaling strategy includes launching a 3000 cm2 prototype biofilm reactor designed to continuously produce chemicals, pushing the boundaries of industrial biotechnology. At the core of the research is the use of the engineered strain Syn6803_capro, which harbors two genes encoding enzymes that catalyze epsilon-caprolactone synthesis. This two-step reaction requires alcohol dehydrogenase (CDH) and Baeyer-Villiger monooxygenase (CHMO) derived from the bacterium Acidovorax sp. strain CHX100. Preliminary work on phototrophic HLM reactors, coupled with their characterization using SEM and their application in spiral-designed bioreactors, provides the experimental foundation and workflow for the 3D-BioCat project. Ultimately, the project aims to transform the production of epsilon-caprolactone through HLMs by using a comprehensive analytical workflow, employing techniques such as confocal laser scanning microscopy, optical coherence tomography, scanning electron microscopy, helium ion microscopy, and both gas and liquid chromatography to track microbial biofilm growth and evaluate catalytic performance within the reactors. Furthermore, we will evaluate if plasmid-based whole-cell catalysts are stable over long reaction times and how plasmid stability and expression profiles are developing in the biofilms. In the future, our new scalable HLMs-based bioreactors should exploit the potential of productive biofilms to continuously produce chemicals and energy carriers.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2494: Productive Biofilm Systems