C1-BOOSTER: Sugar & methanol co-consumption for microbial zero CO2 conversions

• Applicants: Professor Dr. Jan Marienhagen; Dr.-Ing. Lars Regestein; Professor Dr. Sebastian Aljoscha Wahl
• Subject Area: Biological Process Engineering
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 560539272

The C1-BOOSTER challenge: Reaching the sustainability goals defined by the United Nations encourages industry and individuals to find sustainable solutions and reduce greenhouse gas emissions in all sectors. Biotechnological production processes use renewable carbon feedstocks and are expected to drive the transition to a more sustainable, biobased economy with a reduced or even zero CO2 footprint compared to contemporary fossil-based processes. Currently, bioprocesses rely mostly on first- or second-generation feedstocks, which are rich in sugars as these are the preferred substrates of most prevailing industrial microorganisms. Typically, CO2 is released due to an imbalance in the degree of reduction or energy during aerobic or anaerobic bioprocesses. Hence, balancing of energy or redox equivalents by feeding suitable co-substrates could be the key to prevent CO2 emissions. Our solution: Establishment of CO2-free production of fine chemicals using methanol as a co-substrate and electron booster. Why methanol as co-substrate? For the future it is expected that excess renewable energy will be available to generate CO2-derived, reduced carbon substrates at scale. Methanol can be simply obtained by electrochemical reduction of CO2, or synthesis from hydrogen gas and CO2. Methanol is a well soluble, pH neutral, and industrially relevant hosts have a high tolerance to this alcohol. The design of the microbial hosts for the CO2-free production of fine chemicals within this project is based on theoretical calculations, including metabolic network optimization as well as thermodynamic considerations. Based on the calculations, two methanol assimilation pathways are especially beneficial for the integration of methanol in the metabolic network: the ribulose monophosphate and the glycolaldehyde-allose 6-phosphate pathway. In combination of either of the two strategies to assimilate methanol, the so-called non-oxidative glycolysis is essential for eliminating CO2 production. All pathways will be introduced into the industrial workhorse Corynebacterium glutamicum using CRISPER-Cas12a based gene editing. Further improvements of the overall pathway activities will be achieved using advanced laboratory evolution methods. Valuable target products that will be synthesized using this chassis strain are: (1) triacetic acid lactone as a potential platform chemical and (2) polyhydroxybutyrate as relevant bioplastic replacement. In addition to the complex strain engineering, bioprocess control and design will be crucial. The organism has to consume the both substrates (methanol and xylose or glucose) and O2 at very specific ratios to achieve zero CO2 formation. The substrate methanol is volatile, stripping losses have to be minimized by e.g. increased pressure or recirculation of the off-gas. At the same time, high O2-transfer is required.

• DFG Programme: Research Grants
• Co-Investigator: Professor Dr. Jörn Kalinowski