(139b) Metabolic Engineering of Clostridium Ljungdahlii for Production of Fuels and Other Chemicals From Carbon Dioxide Via Microbial Electrosynthesis or with Carbon Monoxide or Hydrogen As the Electron Donor

Clostridium ljungdahlii is an acetogenic microorganism that has the natural ability to use hydrogen as the electron donor to reduce carbon dioxide as its terminal electron acceptor with the production of acetate that is excreted from the cell.  Carbon monoxide can also serve as an electron donor for acetate production.  Under some conditions ethanol and butanediol are produced in addition to acetate, but usually in lesser quantities.  Recent studies in our laboratory have also demonstrated that C. ljungdahlii can directly accept electrons from a negatively poised electrode in order to reduce carbon dioxide to acetate. The ability of microorganisms like C. ljungdahlii to convert carbon dioxide to organic products with electrons derived from an electrode as the driving force is known as microbial electrosynthesis.  Carbon dioxide is reduced to organic products in C. ljungdahlii via the Wood-Ljungdahl pathway.  Acetyl-CoA is a key intermediate in the Wood-Ljungdahl pathway, which offers the possibility of genetically modifying C. ljungdahlii to make a diversity of products more valuable than acetate.  Techniques for genetic manipulation of C. ljungdahlii were developed which permitted deleting genes from, or inserting genes into, the chromosome.  Initial gene manipulation studies focused on the mechanisms for hydrogen uptake and identified a single hydrogenase gene that was required for growth on hydrogen.  This hydrogenase-deficient strain continued to reduce carbon dioxide on negatively poised electrodes, demonstrating that hydrogen was not an intermediate in electron transfer from electrodes to C. ljungdahlii.  Introducing the appropriate genes and/or deleting genes for unwanted pathways made it possible to divert metabolism in C. ljungdahlii away from acetate and ethanol production and toward the production of more desirable products.  A strain for the production of the transportation fuel butanol was developed, as were strains that can produce other multi-carbon organic commodities from carbon dioxide with an electrode or hydrogen serving as the energy source.  Furthermore, these strains could ferment sugars to desired products.  These results demonstrate that C. ljungdahlii is a genetically tractable chassis organism for the production of fuels and other commodities from a diversity of renewable energy sources.