As part of the collaborative project SynergyFuels, three professorships from the TUM-Campus Straubing are responsible for the isobutanol synthesis, namely Professorship Microbial Biotechnology (TUM MIB), Chair of Chemistry of Biogenic Raw Resources (TUM CBR), and Professorship Bioprocess Engineering (TUM BVT).
TUM MIB unlocked and characterized the production of isobutanol from wheat straw hydrolysate with Corynebacterium glutamicum. During the hydrolysis of lignocellulose, different cell toxins including aromates, aldehydes, and aliphatic acids can be formed aside from sugars. We could show that C. glutamicum is robust against various representatives of these inhibitors. Acetate, however, disturbs the energy metabolism, which diminishes the glucose uptake rate. Moreover, we found that a so-far unknown hydrolysate component can entirely disrupt the isobutanol biosynthesis pathway. To optimize the alcohol’s synthesis from wheat straw hydrolysate, we continuously tailor the production strain and test reconditioning the hydrolysate. Moreover, we are developing an antibiotic-independent isobutanol production in a fed-batch process.
So far in the professorship of Bioprocess Engineering (BVT) at TUM Campus Straubing, in-situ product removal (ISPR) methods have been investigated for the removal of isobutanol from fermentation broth. Liquid-liquid extraction has emerged as the most suitable removal method, and several solvents for this purpose have been identified and characterized. Two examples of these solvents include oleyl alcohol and tributyrin. As a proof of concept for in-situ liquid-liquid extraction, isobutanol was extracted from fermentation broth using oleyl alcohol and successfully recovered through distillation. Additionally, a 42 L stainless-steel bioreactor is currently being established at the BVT professorship in preparation for the scale-up of the final process.
At TUM CBR, the main focus is on the biocatalytic production of isobutanol – a branched C4 alcohol with great potential as a future biofuel. In the SynergyFuels concept, isobutanol will also serve as a key intermediate for the production of fleet-suitable OM(B)E fuels. In particular, we are interested in various cell-free multi-enzyme cascades that utilize biogenic resources such as sugar from biological waste and atmospheric CO2 to produce isobutanol in a climate-friendly manner. Previous studies have shown that some of the relevant enzymes exhibit insufficient activities and stabilities, which is problematic given the intended industrial application. Our goal is therefore to improve the isobutanol production of our synthetic biosystems by using enzyme engineering, high-throughput screening methods and cascade optimization.