(495d) Elementary Mode Analysis of Ralstonia Eutropha Metabolism for the Production of Useful Metabolites

Ralstonia eutropha H16 is a Gram-negative, facultative chemolithoautotrophic bacterium with the capability to synthesize many useful metabolites. One of the keys to the organism's lifestyle is its ability to use—alternatively or concomitantly—both organic compounds and molecular H2 as sources of energy. In the absence of environmental O2, R. eutropha H16 can switch to anaerobic respiration; a complete denitrification pathway allows the organism to exploit alternative electron acceptors such as NO3- and NO2-. Correlated with the flexible bioenergetics of the organism is its capacity to shift between heterotrophic and autotrophic growth modes. It fixes CO2 via the Calvin-Benson-Bassham (CBB) cycle and  produces several useful metabolites like Poly(3-hydroxybutyric acid), isobutanol , 2,3-butanediol and ethanol. It can be easily manipulated genetically for the creation of knock out mutations, for integration of genes into the chromosome and it can maintain plasmids with appropriate selection techniques. To quantitatively evaluate the capabilities of the metabolism we have set up a model of the central metabolism that is based on known pathways of the organism. This model includes heterotrophic as well chemolithoautotrophic growth modes.   The metabolic model consists of 21 reversible reactions, 51 irreversible reactions, 56 internal metabolites and 16 external metabolites exchanged through the cell membrane. Elementary Mode analysis on this model identified 55,585 modes for the litho-autotrophic growth mode and 61521 modes for the heterotrophic mode on fructose as carbon and energy source.  We have used the results from this analysis to predict key genetic alterations in the metabolism that would direct the metabolic flux towards metabolites of interest including basic and advanced bio fuels as well as Poly(3-hydroxybutyric acid) and other secondary metabolites.  The experimental realization of predicted metabolic networks is currently underway.