Bioprinting of Nanocellulose Hydrogels for Photobiocatalysis in Continuous Flow Systems

Photosynthetic microorganisms are capable of oxygenic photosynthesis delivering both, oxygen and cofactors to drive enzymatic redox reactions. However, their dependence on visible light limits the tolerable cell densities to achieve high reaction rates. Immobilising cells within a matrix often increases biocatalyst productivity while allowing facile retainment, but also creates mass transfer limitations across the solid-liquid interface. Herein, we have immobilised recombinant cyanobacteria in 3D-printed hydrogels of varying geometries. The hydrogels were 3D-printed via extrusion into flow reactors using a combination of biocompatible nanofibrillated cellulose and alginate to alleviate light and mass transfer limitations. The reactors exhibit high mechanical stability, improved flow and mass transfer characteristics, increased specific surface area and longer retention times. This allowed upscaling of the bioreactor leading to higher product formation rates. Whole cells of the cyanobacterium Synechocystis sp. PCC 6803 engineered to express the gene of the ene-reductase YqjM were employed to reduce prochiral 2-methylmaleimide to (R)-2-methylsuccinimide with high enantiomeric purity (>99%) and a space-time yield (STY) of 0.28 g L-1 h-1. This highly atom-efficient chemical process (88%) using only water to provide electrons for NADPH regeneration can potentially be operated in extended periods to reduce wastewater associated with cell cultivation. Overall, 3D-printing of photosynthetic microorganisms embedded in a hydrogel matrix holds significant promise for advancing the development of whole-cell solid state photosynthetic cell factories. These are important steps towards improved reactor designs and higher efficiencies to improve crucial redox biotransformations.