Periodic Reporting for period 1 - TransCO2 (Electricity-Driven Enzymatic Cascades to Transform CO2 to C2+ Chemicals)
Periodo di rendicontazione: 2023-08-01 al 2025-07-31
The increasing level of carbon dioxide (CO2) in the atmosphere presents a critical factor for climate change and action must be taken urgently to minimise its impact. In addition to the necessary reduction of CO2 emissions, there are efforts to use CO2 as a carbon source in the chemical industry for its conversion into valuable chemicals, which is an advantageous strategy to reduce CO2 emissions and provide a sustainable and cheap source of raw materials to help combat raw material scarcity. Electrochemistry allows for the utilization of an electrical input to drive chemical reactions. Multi-enzyme cascades have set the benchmarks for making complex molecules from CO2 with high control on regio-, chemo and stereoselectivity. Interfacing these CO2 reducing enzyme/enzymatic cascades with electrocatalysis would be ideal to directly power product generation from CO2 with renewable electricity. This was achieved for a few enzymatic cascades; however, these proof-of-concept demonstrations are far from practical use due to lack of efficient method to guide the rational design of these complex multi-enzymes cascades on electrodes, resulting in high costs and low yields. Within this Marie Curie Postdoctoral fellowship project (TransCO2), my overarching aim is to apply quantitative analysis and rational assembly of enzymatic cascades to enable a breakthrough in bioelectrocatalytic-technology to transform carbon dioxide (CO2) directly into high value mevalonate (C5) at high isolated yields, using electricity as energy source. Specifically, CO2 is firstly converted to formate by the enzyme Formate dehydrogenase (FDH), which is then further converted to the key intermediate acetyl-CoA via 4 steps enzymatic conversions. Then the acetyl-CoA can be applied to produce various valuable chemicals, like bioplastics, biofuel and natural compounds. In this project, I choose to produce the mevalonate as a case of study. The whole system will function in an electrochemical cell to make use of electricity to directly power FDH, but also for other two enzymes (FalDH and Hmgr) in the cascade. I will build a kinetic model for this enzymatic cascade and implement it to guide the design and optimisation towards highest efficiency with minimal utilization of expensive cofactors. The overall technology developed in TransCO2will be a breakthrough for efficient and powerful bioproduction technology for the production of high-value chemicals from CO2, enabling the large-scale use of CO2 utilisation. TransCO2 will output a powerful and general biomanufacturing technology by integrating the bioelectrocatalysis and multiple enzymatic cascade conversions, which will significantly promote the research on designing novel sustainable synthetic systems. For example, regarding current researches on cell free synthetic enzymatic cascades, the popular strategy is to reconstitute the glycolysis pathway (11 steps) to firstly convert glucose to the key intermediate acetyl-CoA, which is then applied to produce various valuable chemicals. TransCO2 will provide a more efficient pathway, only 5 steps are sufficient to obtain acetyl-CoA and the substrate CO2 is more easy-accessible and sustainable than glucose, both of which will make this system cost-effective for industrial applications. This will be further consolidated by introducing the advanced mathematic modelling strategy to reach their optima, which can be applied to other existing CO2 transformation systems, enhancing their industrial potentials. In addition, the inherent open environment of such systems facilitates the engineering, leading to a high generality. For example, many other valuable chemicals, including terpenes and polyketides, isobutanol and isoprene, can be easily produced from the key intermediate acetyl-CoA, simply by adding related enzymes. Furthermore, this modern technology will highly benefit to long-term crewed missions on Mars, of which a major limiting factor is the cost of launching goods into space as stated by NASA. Transformation of in situ source (N2, CO2, water) to fuel, food, biomaterials and medicine, will minimize the quantity and frequency of resupply missions that will otherwise be required due to limited food and pharmaceutical shelf-life.
1. Expression in Ecoli of enzyme FDH and FaldDH and purification
We did a screen on various FDH and FaldDH reported in the literatures at first, the formate dehydrogenase from Clostridium ljungdahlii and formaldehyde dehydrogenase from Burkholderia multivorans are chosen. After several rounds of optimisation on the expression and purification conditions, we managed to obtain these two enzymes with high purifity. And their acitivity and kinetic parameters are determined based on UV measurements.
2. Development of novel redox metrix to wire enzymes on electrode
To obtain high catalytic currencies from the enzymes on electrodes, 3D immobolisation of enzymes on electrodes are reuqired and the metrix is capable to mediate electrons between enzymes and electrodes. Using Hydrogenase as a model enzyme, we certify we have developed a novel reodx metrix which could stably wire enzymes on electrodes for days under continous operations.
We did a screen on various FDH and FaldDH reported in the literatures at first, the formate dehydrogenase from Clostridium ljungdahlii and formaldehyde dehydrogenase from Burkholderia multivorans are chosen. After several rounds of optimisation on the expression and purification conditions, we managed to obtain these two enzymes with high purifity. And their acitivity and kinetic parameters are determined based on UV measurements.
2. Development of novel redox metrix to wire enzymes on electrode
To obtain high catalytic currencies from the enzymes on electrodes, 3D immobolisation of enzymes on electrodes are reuqired and the metrix is capable to mediate electrons between enzymes and electrodes. Using Hydrogenase as a model enzyme, we certify we have developed a novel reodx metrix which could stably wire enzymes on electrodes for days under continous operations.
Impacts:
Here we have managed to express and purify a new mental dependent FDH in E coil, which shows high activity in CO2 reduction to formate. This work will directly impact contemporary societal challenges - climate change and global warming, resulting from the excessive emissions of greenhouse gases. To circumvent these severe consequences and avoid the point of no return, the Paris Agreement on ‘2-degree scenario’, aiming to keep warming at or below 1.5 oC, has been made in 2015 and accepted globally, which requires reducing the emission of greenhouse gas including CO2. This work will make an important step towards achieving this goal by presenting an efficient system to fix CO2, by making the existing systems more industrially relevant and by introducing a general technology to design more CO2 transformation pathways. In addition, This work creates a way to utilize CO2 as cheap, nontoxic, and naturally abundant carbon source instead of reagents derived from fossil-fuels. This will help decrease the dependence on fossil-fuels, which in return decreasing the emission of CO2. In summary, This work represents a double gain trial and is highly attractive from both ecological and economic viewpoints.
In addition, we have develop a new redox metrix to wire enzymes on electrodes. This will provide a new method to make use of green electricity to power enzymatic synthesis.
Here we have managed to express and purify a new mental dependent FDH in E coil, which shows high activity in CO2 reduction to formate. This work will directly impact contemporary societal challenges - climate change and global warming, resulting from the excessive emissions of greenhouse gases. To circumvent these severe consequences and avoid the point of no return, the Paris Agreement on ‘2-degree scenario’, aiming to keep warming at or below 1.5 oC, has been made in 2015 and accepted globally, which requires reducing the emission of greenhouse gas including CO2. This work will make an important step towards achieving this goal by presenting an efficient system to fix CO2, by making the existing systems more industrially relevant and by introducing a general technology to design more CO2 transformation pathways. In addition, This work creates a way to utilize CO2 as cheap, nontoxic, and naturally abundant carbon source instead of reagents derived from fossil-fuels. This will help decrease the dependence on fossil-fuels, which in return decreasing the emission of CO2. In summary, This work represents a double gain trial and is highly attractive from both ecological and economic viewpoints.
In addition, we have develop a new redox metrix to wire enzymes on electrodes. This will provide a new method to make use of green electricity to power enzymatic synthesis.