Building a light driven synthetic carbon dioxide fixation cycle within microdroplets
Cells are highly integrated biological systems that perform complex tasks. These self –sustained compartments exist thermodynamically out-of-equilibrium with the environment and require a constant influx of energy to drive the internal metabolism and prevent decay to equilibrium. Photosynthetic auto...
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|Cells are highly integrated biological systems that perform complex tasks. These self –sustained compartments exist thermodynamically out-of-equilibrium with the environment and require a constant influx of energy to drive the internal metabolism and prevent decay to equilibrium. Photosynthetic autotrophic organisms convert light into chemical energy, which is the driving force for the transformation of inorganic carbon into organic compounds. Ultimately, the photosynthetic conversion of light energy proceeds through membrane bound protein complexes, which generate the energy-rich chemical cofactors adenosine triphosphate (ATP) and reduced nicotinamide adenine dinucleotide phosphate (NADPH). ATP and NADPH are subsequently used to fuel metabolic processes, in particular the fixation of carbon dioxide (CO2) through the Calvin-Benson-Bassham (CBB) cycle. So far, efforts to create an artificial cell or organelle that mimics autotrophic photosynthesis have not succeeded in linking light harvesting and carbon fixation at the micron scale. In this work, microfluidics and synthetic biology were combined in an attempt to develop and optimize a functional mimic of a chloroplast in a mostly bottom-up fashion.
In this work, a photosynthetic energy module was developed based on thylakoid membranes of spinach chloroplasts, its function optimized, and then used to power different enzymatic reactions and complex metabolic networks by light. Microfluidic-based encapsulation of the photosynthetic energy module generated cell-sized droplets that can be equipped with enzymes, energized by light and analyzed for catalytic function in multiplex and real-time. The activity of the micro-droplets can be programmed and controlled by adjusting internal compositions (e.g. thylakoid membranes and enzyme concentrations) as well as using light as an external trigger.
Coupling this photosynthetic energy module with a 17-enzyme, new-to-nature CO2-fixation cycle, created a structural and functional mimic of a chloroplast that continuously converts CO2 into the organic compound glycolate. In essence, natural and synthetic parts have been combined to drive anabolic reactions by light at a micron scale. This platform represents a basis useful for multiple applications in both top-down and bottom-up synthetic biology approaches, while also signifying another step on the way towards creating functional mimics of living cells.