(602f) Enhancement of H2 and CO2 Uptake for the Production of Biodiesel in Ralstonia Eutropha

Ralstonia eutropha fixes CO₂ through the Calvin-Benson-Bassham (CBB) cycle using electrons and energy obtained from the oxidation of H₂.  Producing biodiesel-equivalent electrofuel from renewable CO₂ and H₂ has immense potential, especially if the fuel is compatible with the existing fuel infrastructure.  This research addressed enhanced H₂ and CO₂ substrate utilization by focusing on two strategies: (1) optimizing transcriptional regulations to afford over-expression of Ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), the enzyme responsible for assimilation of CO₂ into the CBB cycle; and (2) hydrogenase over-expression by introduction of additional copies of genes encoding a membrane-bound hydrogenase (MBH), a soluble hydrogenase (SH), and their maturation machinery to enhance oxidation of H₂ to generate NAD(P)H and ATP, both are required for CO₂ fixation.  Incorporation of these strategies into a single production R. eutropha strain had resulted in 6-fold improvement in CO₂ fixation and 3-fold improvement in H₂ uptake, in vitro, with the overarching goal of providing abundant reducing equivalents towards the economic production of biodiesel in R. eutropha.